text:
- Research article
- Open Access
Detection of autoreactive CD4 T cells using major histocompatibility complex class II dextramers
© Massilamany et al; licensee BioMed Central Ltd. 2011
- Received: 28 April 2011
- Accepted: 18 July 2011
- Published: 18 July 2011
Tetramers are useful tools to enumerate the frequencies of antigen-specific T cells. However, unlike CD8 T cells, CD4 T cells - especially self-reactive cells - are challenging to detect with major histocompatibility complex (MHC) class II tetramers because of low frequencies and low affinities of their T cell receptors to MHC-peptide complexes. Here, we report the use of fluorescent multimers, designated MHC dextramers that contain a large number of peptide-MHC complexes per reagent.
The utility of MHC dextramers was evaluated in three autoimmune disease models: 1) proteolipid protein (PLP) 139-151-induced experimental autoimmune encephalomyelitis in SJL/J (H-2s) mice; 2) myelin oligodendrocyte glycoprotein (MOG) 35-55-induced experimental autoimmune encephalomyelitis in C57Bl/6 (H-2b) mice; and 3) cardiac myosin heavy chain (Myhc)-α 334-352-induced experimental autoimmune myocarditis in A/J (H-2a) mice. Flow cytometrically, we demonstrate that IAs/PLP 139-151, IAb/MOG 35-55 and IAk/Myhc-α 334-352 dextramers detect the antigen-sensitized cells with specificity, and with a detection sensitivity significantly higher than that achieved with conventional tetramers. Furthermore, we show that binding of dextramers, but not tetramers, is less dependent on the activation status of cells, permitting enumeration of antigen-specific cells ex vivo.
The data suggest that MHC dextramers are useful tools to track the generation and functionalities of self-reactive CD4 cells in various experimental systems.
- Antigen-specific CD4 cells
- Central nervous system
- Dextramers
- Experimental autoimmune encephalomyelitis
- Experimental autoimmune myocarditis
- Heart
- Major histocompatibility complex class II
- Myelin oligodendrocyte glycoprotein
- Cardiac myosin heavy chain-alpha
- Proteolipid protein
- Tetramers
Traditionally, limiting dilution analysis and cytokine ELISPOT assays have been used to enumerate the frequencies of antigen-specific cells, but low specificity and the tedious nature of these assays limit their use for routine applications [1–3]. To overcome these limitations, tetramer technology has been developed [4]. This approach involves derivation of fluorescent dye-labeled tetramerized complexes containing peptide-assembled major histocompatibility complex (MHC) molecules. The use of tetramers permits easy and rapid detection of antigen-sensitized T cells at a single cell level by flow cytometry (FC) [4, 5]. Additionally, tetramer reagents can be used in conjunction with antibodies to phenotypically characterize the generation and expansion of antigen-specific cells during ensuing immune responses and to study their functionalities by cell sorting.
Unlike with MHC class I tetramers, direct enumeration of antigen-specific cells - in particular, rare and low-affinity autoreactive CD4 T cells - with MHC class II tetramers has been difficult. Under natural conditions, interaction between T cells and antigen-presenting cells involves engagement of MHC molecules with multiple T cell receptors (TCRs), but their binding strength is low. Hence, their stability requires interaction between various adhesion molecules. Tetramers are artificially created peptide-assembled MHC complexes, and their interactions with TCRs occur in the absence of accessory molecules, a limitation that exists with both class I and class II tetramers. In spite of this limitation, class I tetramers can generally bind CD8 T cells, but require participation of a CD8 coreceptor [6]. By comparison, participation of a CD4 coreceptor is not critical for class II tetramers to bind to CD4 cells [7, 8]. This differential requirement of coreceptors may, in part, explain differences in the abilities of tetramers to bind respective T cell populations.
It is generally accepted that class II tetramers preferentially bind activated CD4 cells expressing high amounts of CD4 and CD25 [9–12]. We and others had previously proposed that activation might result in reorientation and configuration of TCR and accessory molecules, leading to enhanced avidity of TCR-MHC binding [11, 13, 14], thus restricting the utility of the reagents for direct enumeration of the frequencies of antigen-sensitized cells ex vivo. Recently, MHC dextramers bearing dextran backbones assembled with peptide-tethered MHC class I molecules have been shown to detect antigen-specific CD8 T cells [15, 16]. Our studies involve the use of a streptavidin (SA)-dextramer conjugate of 270 kDa, which has been validated to detect tumor- and virus-specific cells in various systems [15, 17–19]. To address the utility of dextramers, we used three autoimmune disease models: 1) proteolipid protein (PLP) 139-151-induced experimental autoimmune encephalomyelitis (EAE) in SJL/J (H-2s) mice; 2) myelin oligodendrocyte glycoprotein (MOG) 35-55-induced EAE in C57Bl/6 (H-2b) mice; and 3) cardiac myosin heavy chain (Myhc)-α 334-352-induced experimental autoimmune myocarditis (EAM) in A/J (H-2a) mice. We demonstrated that MHC class II dextramers detect self-reactive CD4 T cells with specificity and to a greater sensitivity than conventional tetramers. We showed that binding of dextramers, but not tetramers, is less dependent on activation status of the cells, thus permitting the reagents to determine the frequencies of antigen-specific cells ex vivo in immunized mice.
Four-to-six-week-old female SJL/J (H-2s) and C57Bl/6 (H-2b) mice and six-to-eight-week-old A/J (H-2a) mice were procured from the Jackson Laboratory (Bar Harbor, ME). The mice were maintained according to the animal protocol guidelines of the University of Nebraska-Lincoln, Lincoln, NE.
Peptide synthesis and immunization procedures
PLP 139-151 (HSLGKWLGHPDKF), MOG 35-55 (MEVGWYRSPFSRVVHLYRNGK) and Myhc-α 334-352 (DSAFDVLSFTAEEKAGVYK) were synthesized on 9-fluorenylmethyloxycarbonyl chemistry (Neopeptide, Cambridge, MA). All peptides were HPLC-purified (>90%), identity was confirmed by mass spectroscopy, and the peptides were dissolved in 1 × PBS prior to use. To generate primary T cell cultures for PLP 139-151 and MOG 35-55, SJL and C57Bl/6 mice, respectively, were immunized with the above peptides emulsified in complete Freund's adjuvant (CFA) subcutaneously (s.c.) in multiple sites in the flank and sternal regions (100 μg/mouse) [5, 20]. For immunization of A/J mice with Myhc-α 334-352, the peptide was emulsified in CFA supplemented with Mycobacterium tuberculosis (M. tb; Difco Laboratories, Detroit, MI) extract to a final concentration of 5 mg/ml and injected s.c. in the flank, foot pads and sternal regions (100 μg/mouse). For disease induction, however, regardless of peptides used, emulsions were prepared in CFA containing M. tb extract to a final concentration of 5 mg/ml. To induce EAE with PLP 139-151 or MOG 35-55, peptide emulsions were given as a single dose [11, 20], whereas for EAM induction, two doses of emulsions containing Myhc-α 334-352 were administered with an interval of one week [21, 22]. Nonetheless, all animals involved in the disease-induced protocols received pertussis toxin (List Biological Laboratories, Campbell, CA) intraperitoneally on day 0 and day 2 after the first immunization (PLP 139-151 and Myhc-α 334-352, 100 ng/mouse; MOG 35-55, 200 ng/mouse) [5, 20, 21]. SJL and C57Bl/6 mice were monitored for the appearance of clinical signs of EAE and scored as described [23, 24]: 0 - healthy; 1 - limp tail or hind limb weakness but not both; 2 - limp tail and hind limb weakness; 3 - partial paralysis of hind limbs; 4 - complete paralysis of hind limbs; 5 - moribund or dead.
Generation of antigen-specific primary T cell cultures
Ten days after immunizations with peptides, mice were sacrificed, and the maxillary, mandibular, axillary, inguinal and popliteal draining lymph nodes (LN) were collected. Single cell suspensions were prepared after lysing the erythrocytes with 1 × ammonium chloride potassium buffer (Lonza, Walkersville, MD). Lymph node cells (LNC) were stimulated with the peptides (PLP 139-151, 20 μg/ml; MOG 35-55 and Myhc-α 334-352, 50 μg/ml) at a density of 5 × 106 cells/ml for two days in RPMI medium supplemented with 10% fetal bovine serum (FBS), 1 mM sodium pyruvate, 4 mM L-glutamine, 1 × each of non-essential amino acids and vitamin mixture, 100 U/ml penicillin-streptomycin and 50 μg/ml gentamicin (Lonza). After two days, cultures were supplemented with the above medium containing interleukin (IL)-2 (eBioscience, San Diego, CA), hereafter termed growth medium. Viable lymphoblasts were harvested two to three days later by Ficoll-Hypaque density gradient centrifugation, and the cultures were maintained in growth medium.
Isolation of mononuclear cells (MNC) from CNS tissues and hearts
After euthanization, mice were perfused by injecting 10 ml of 1 × cold PBS into the left ventricle of the hearts; brains were harvested by blunt dissection; and spinal cords were flushed out using 1 × cold PBS. The tissues were homogenized and digested with type IV collagenase (400 U/ml; Worthington, Lakewood, NJ) at 37°C for 1 hour, and MNC were harvested by percoll density gradient centrifugation (70%/30%) as described previously [25]. To harvest MNC from hearts, animals were first perfused as above, and hearts were collected and minced. The tissue suspensions were then incubated with a buffer containing 1 × PBS, collagenase and 2% FBS at 37°C for 15 min in a shaker with continuous agitation. The digested tissues were passed through an 18G needle and incubated for an additional 10 minutes; the cell pellets were obtained by centrifugation and resuspended in 40% buffered percoll. Cell suspensions were overlaid with 75% percoll and centrifuged; the interphase representing MNC was then collected and washed [26, 27].
Derivation of IAs, IAb and IAk tetramers and dextramers
IAs/PLP 139-151 and IAs/Theiler's murine encephalomyelitis virus (TMEV) viral capsid protein2 70-86, IAb/MOG 35-55, and IAk/Myhc-α 334-352 and IAk/Bovine ribonuclease (RNase) 43-56 tetramers were generated as previously described [5, 11, 25, 28]. While we generated the IAk constructs [11, 29], the IAs and IAb constructs were kindly gifted by Dr. Vijay Kuchroo, Harvard University, Boston, MA. Briefly, α and β constructs for each IA allele containing the sequences of the respective peptides were expressed in a Baculovirus system using Sf9 insect cells (Invitrogen, Carlsbad, CA) and soluble MHC molecules were obtained [5, 11]. IAs, IAb and IAk monomers were purified on antibody columns prepared using MKS4, M5114, and 10-2.16 (Bio × Cell, West Lebanon, NH), respectively and the protein yield generally ranged from 0.5 mg to 1 mg/L [5, 11, 25, 28]. After concentrating, the soluble MHC proteins were biotinylated using biotin protein ligase at an optimized concentration of 25 μg/10 nmol of substrate as recommended by the manufacturer (Avidity, Denver, CO). The biotinylated proteins were then incubated with SA conjugated with a fluorescent dye - fluorescein isothiocyanate (FITC), phycoerythrin (PE) or allophycocyanin (APC) - at a 4:1 ratio for one hour on ice. The reagents thus prepared are referred to as tetramers. To prepare dextramers, biotinylated soluble monomers of all three IA alleles (IAs, IAb and IAk) containing the peptides were coupled to activated dextran backbones (kindly provided by Immudex Aps, Copenhagen, Denmark) at various molar ratios in 1 × Tris Hcl 0.05 M, pH 7.2 for 30 minutes at room temperature (RT) and the preparation of fluorochrome-labeled dextran backbone has been previously described [16]. Fluorochrome-labeled dextramers were prepared by mixing biotinylated IA monomers with dextran molecules. For example, dextramer-PE reagents were prepared by mixing 1.6 × 10-11 moles of dextran-PE molecules with 3.17 × 10-10 moles of IAs monomers which resulted a molar ratio of 1:20. All the reagents were aliquoted and stored at -80°C or 4°C until further use.
Tetramer or dextramer staining
(a) Cells activated in vitro
To enumerate the frequencies of tetramer (tet+) or dextramer (dext+) cells, viable lymphoblasts harvested from cultures prepared from immunized SJL, C57Bl/6 or A/J mice were used for tet or dext staining as described previously [5, 30]. Briefly, cells were stained with tetramers or dextramers in growth medium, pH 7.6, containing 2.5% FBS at RT for up to 3 hours, followed by staining with anti-CD4 (eBioscience, San Diego, CA) and 7-aminoactinomycin D (7-AAD; Invitrogen). After washing, cells were analyzed by flow cytometry (FC; FACSCalibur, BD Biosciences, San Diego, CA). Percentages of tet+ or dext+ cells were then determined in the live (7-AAD-) CD4 subset using Flow Jo software (Tree Star, Ashland, OR). In some experiments, antigen-sensitized cells were rested for two weeks in growth medium and treated with or without neuraminidase (NASE) (0.7 U/ml) (Type X from Clostridium perfringens; Sigma-Aldrich, St. Louis, MO) prior to tet or dext staining [11].
(b) Ex vivo staining
LNC were obtained from a pool of draining LN harvested from groups of mice immunized with various peptides. CD4 cells were enriched from LNC to a purity of more than 95% by negative selection based on magnetic separation using IMAG as recommended by the manufacturer (BD Biosciences). After treating the cells with or without NASE as above, cells were washed in 5 ml of 1 × PBS; incubated with tetramers or dextramers for two hours at RT in growth medium, pH 7.6, containing FBS (2.5%). Cells were washed twice with 5 ml of 1 × PBS and stained with anti-CD4, anti-CD25, anti-CD44 (all from eBioscience) and 7-AAD and analyzed by FC. The percentages of tet+ or dext+ cells were determined in the live (7-AAD-) cells corresponding to CD4, CD25 and CD44 subsets. To stain MNC obtained from CNS tissues, cells were incubated with tetramers or dextramers directly without NASE treatment, and the percentages of tet+ or dext+ cells were determined as described above.
Sorting of tet+ or dext+ cells by flow cytometry
LNC isolated from immunized mice were stimulated with PLP 139-151 at a density of 5 × 106 cells/ml and viable lymphoblasts were harvested on day 6 poststimulation and stained with tetramers or dextramers for two hours at RT. After washing, cells were stained with anti-CD4 and 7-AAD; the tet-CD4+ and dext-CD4+ cells were sorted by FC (FACSAria, BD Biosciences, San Jose, CA), and each fraction was divided into two aliquots. One aliquot of cells was immediately stained with APC-labeled PLP 139-151 or control dextramers for two hours, followed by 7-AAD. After acquiring the data by FC, dext+ cells were analyzed in the live (7-AAD-) subset. The second aliquot (0.75 × 106 cells/ml) was rested in growth medium for two weeks and stimulated with PLP 139-151 (10 μg/ml) in the presence of irradiated antigen-presenting cells for two days; growth medium was then supplemented. Viable lymphoblasts were harvested two days later, and on day 6 poststimulation, PLP 139-151 or control tet or dext staining was performed followed by anti-CD4 and 7-AAD and the percentages of live tet+ or dext+ cells were enumerated as above.
Differences between specific and control tet or dext and also between the groups were analyzed by student's t. P ≤ 0.05 values were considered significant.
Optimization of reaction conditions for dextramer staining
FITC-labeled dextramers but not tetramers detect antigen-reactive cells
In spite of high sensitivity, dextramers fail to detect all antigen-reactive cells
Activation dependency is less stringent for dextramer binding
Dextramers are useful tools to enumerate the frequencies of antigen-specific cells ex vivo
Using three different murine autoimmune disease models, we have demonstrated that MHC class II dextramers permit ex vivo enumeration of self-reactive CD4 T cells with specificity. Tetramers, but not monomers or dimers, are useful in detecting CD4 T cells by FC [4, 12, 33]. In spite of this technical advancement, detection of autoreactive CD4 cells is not straightforward because of their rarity in the peripheral repertoires, and their detection often requires in vitro expansion and/or enrichment [33]. In this scenario, however, while antigen specificity of cells can still be confirmed, accurate estimation of their frequencies can be prone to error because of the need to activate cells prior to tetramer staining. Structurally, dextramers contain dextran backbones, which are polymers of glucose molecules attached through 1-6 and 1-3 linkages [15]. Each dextran molecule carries multiple moieties of SA to which biotinylated peptide-tethered MHC molecules can be assembled [15]. Therefore, MHC dextramers can yield MHC-peptide complexes of large molecular weight, allowing them to engage multiple TCRs - more than that could be achieved with tetramers. To address this hypothesis, we created MHC dextramers for three mouse IA alleles, IAs, IAb and IAk for PLP 139-151, MOG 35-55 and Myhc-α 334-352, respectively, and showed that the reagents detect corresponding antigen-sensitized cells with specificity and also to a higher sensitivity than conventional tetramers.
We optimized the conditions for dextramer staining using PLP-reactive cells generated from SJL mice immunized with PLP 139-151. Essentially, the conditions previously defined for conventional tetramers were also optimum for dextramers, and the staining was performed in growth medium supplemented with 2.5% FBS, pH 7.6 [5, 11]. However, we noted some important advantages with dextramers, including enhanced sensitivity, shortened staining duration and the ability to create dextramers with a low amount of MHC/peptide complexes. We found that dextramers prepared at a 1:20 molar ratio of SA-dextran molecules to biotinylated MHC proteins were optimum to detect PLP-reactive cells, and their detection sensitivity was ~4-fold higher than that of tetramers. Importantly, the use of dextramers allowed us to capture antigen-specific cells by FC unambiguously as observed by an increase in their mean fluorescent intensities in relation to tetramers (Figure 1). This is an important consideration in flow cytometric analysis, especially when FITC-labeled reagents are used. In support of this notion, PLP 139-151 dextramers, but not tetramers, labeled with FITC detect nearly equivalent percentages of PLP-reactive cells similar to percentages obtained with PE- or APC-labeled dextramers. Furthermore, the enhanced detection of antigen-reactive cells could be achieved as early as 30 minutes postincubation with dextramers, whereas with tetramers, the staining intensity was only marginally improved even when extending the incubation time up to 3 hours. On a reaction basis, we calculated the amount of MHC protein required to prepare tetramers to be 2- and 4-fold higher than the amount of protein required to make dextramers labeled with PE and APC, respectively. The marginal reduction in the staining intensity obtained with FITC-dextramers was expected because signals generated from FITC are weaker than other fluorescent dyes. The fact that FITC-labeled dextramers, but not tetramers, bound PLP-reactive cells provides opportunities to include this dye in the FC-based multicolor analysis of antigen-specific T cell populations.
We believe that the higher detection sensitivity of dextramers is due to the presence of multiple SA moieties, allowing the formation of large clusters of MHC-peptide complexes, which can interact with multiple TCRs, thus enhancing the avidity of their interactions [15, 33]. Conversely, each tetramer molecule contains a single SA moiety to which only up to four MHC-peptide complexes can bind, limiting the availability of MHC-peptide complexes to engage with multiple TCRs. As a result, tetramers might bind T cells bearing high- but not low-affinity TCRs. This appears to be the case because a proportion of tet- cells sorted by FC could still be detected by dextramers (Figure 4A). Nevertheless, it should be noted that dextramers do not detect all antigen-sensitized cells, as indicated by the fraction of PLP-reactive cells previously negative for PLP dextramers that can become detectable after a second restimulation with PLP 139-151 (Figure 4B). Presently the significance of these cell populations is not known with respect to their TCR-affinities or functionalities, such as cytokine synthesis or encephalitogenicity. Alternatively, failure to detect all antigen-reactive cells with dextramers might be directly related to the density of TCRs expressed on the surface of T cells such that low TCR-expressed cells may escape detection. From a technical point of view, formation of stable MHC-peptide-TCR interactions with dextramers can help cells withstand harsh steps such as centrifugation and washing, thereby enhancing specificity by minimizing the background staining. Overall, the fact that dextramers permit quick determination of antigen-specific cells with enhanced sensitivity and because their creation requires less starting material, they have proven to be superior to conventional tetramers.
One of the limitations of class II tetramer binding is activation dependency [9–12]. Various strategies have been employed to enhance the detection sensitivity of class II tetramers by promoting interactions with multiple TCRs. These include antibody-mediated TCR clustering, MHC-peptide-coated liposomes and immunoglobulin molecules [14, 34–36]. We had previously shown that, upon resting, antigen-sensitized cells that have expanded in vitro become undetectable with tetramers, but can be detected through NASE treatment prior to staining [11]. We asked whether the use of dextramers can overcome as the need for NASE treatment or activation. Expectedly, dextramers could bind rested cells in the absence of NASE treatment to a significantly greater proportion than that achieved with tetramers (Figure 5). The data suggest that activation dependency is not a critical factor for dextramer staining, and we verified this phenomenon further by ex vivo enumeration of PLP-specific cells in SJL mice immunized with PLP 139-151 (Figure 6). While, similar results were obtained with IAb/MOG 35-55 and IAk/Myhc-α 334-352 dextramers, NASE treatment added the advantage of enhancing their detection sensitivity as compared with tetramers (Figures 7 and 8). The data prove that MHC dextramers are useful tools for enumerating the frequencies of antigen-specific T cells ex vivo in various experimental systems.
We have shown that dextramers are superior to tetramers in detecting autoreactive cells, which otherwise can be technically challenging. While activation dependency is less important for dextramer binding, detection sensitivity can be enhanced by NASE treatment without compromising the specificity. Although dextramers cannot detect all cells in spite of being antigen specific, their greater detection sensitivity makes them first-choice reagents over conventional tetramers. Additionally, the inherent variation in the detection abilities of these reagents is to be considered while interpreting the results because frequencies of antigen-specific cells identified by each reagent are expected to be different.
We thank Immudex, Denmark for providing dextramer reagents (contact person: Tina Jakobsen, email@example.com), and Dr. Jean-Jack Riethoven for statistical analysis. This work was co-funded by the American Heart Association and the Children's Cardiomyopathy Foundation (SDG2462390204001).
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- Nepom GT: MHC multimers: expanding the clinical toolkit. Clin Immunol. 2003, 106: 1-4. 10.1016/S1521-6616(02)00014-1.PubMedView ArticleGoogle Scholar
- Falta MT, Fontenot AP, Rosloniec EF, Crawford F, Roark CL, Bill J, Marrack P, Kappler J, Kotzin BL: Class II major histocompatibility complex-peptide tetramer staining in relation to functional avidity and T cell receptor diversity in the mouse CD4(+) T cell response to a rheumatoid arthritis-associated antigen. Arthritis Rheum. 2005, 52: 1885-1896. 10.1002/art.21098.PubMedView ArticleGoogle Scholar
- Hugues S, Malherbe L, Filippi C, Glaichenhaus N: Generation and use of alternative multimers of peptide/MHC complexes. J Immunol Methods. 2002, 268: 83-92. 10.1016/S0022-1759(02)00202-8.PubMedView ArticleGoogle Scholar
- Kotzin BL, Falta MT, Crawford F, Rosloniec EF, Bill J, Marrack P, Kappler J: Use of soluble peptide-DR4 tetramers to detect synovial T cells specific for cartilage antigens in patients with rheumatoid arthritis. Proc Natl Acad Sci USA. 2000, 97: 291-296. 10.1073/pnas.97.1.291.PubMedPubMed CentralView ArticleGoogle Scholar
This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.


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No matter what colors you have in your kitchen you will find something to match with this awesome kitchen gear sale with prices that start t just $7. They have great deals on some really great brands!! Just head on over to One King's Lane. If you have not shopped with One King's Lane yet you will get a FREE $15 credit that can be used on a purchase of $30 or more. Get your FREE $15 credit and then check out the sale HERE
They have the Reston Lloyd Colanders for only $15! I have this one in red and I LOVE mine!! They have is in pink, yellow, blue, green, and you can pick from the 1qt, 1.5qt or the 3qt one 🙂 If you buy 2 and use your FREE $15 Credit they are only $7.50 each!! (reg $22).
They also have lots of Kalorik Appliances on sale! Which I love that you can match just about any kitchen with the colored appliances!
Check out the sale HERE

subset: commoncrawl


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Charter Croazia, Noleggio Barche in Croazia
Najam plovila u Hrvatskoj, Hrvatski
Yacht charter Croatia, boat rental and yacht hire in Croatia
Yachtcharter Kroatien, Njemački
When planning charter of the boat make sure that you do it in advance - the advantage of early booking lies in the fact that you will get the wished boat model for a reasonable and acceptable price.
If you can pack your bags fast - our last minute charter section is the perfect place for you! Last minute deal gathers in one place all the boats that are left available for a close departure dates and that can be chartered for a pretty convenient and attractive price. Take the advantage of the unbelievably low charter rates and save up to 30% on the regular charter prices!
You are looking for the new boat model? One-way charter? Regatta deals? Special offers, seasonal offers and all other special charter arrangements this way can be found in one place and are at your disposal for perfect sea summer holiday. Book the wished boat through our SPECIAL DEAL offer and save money by cutting down your charter expences – the fun sea activities are waiting for you!
In our Best buy section we have selected all the boats that are offering best value for money. The best price range, the best equipment or the best sailing area are just some criteria for which the boat deserves to be in this category. Pick some of the boats from this list and find the boat that will provide you the unforgettable summer experience!
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This site is registered with Yachts.com and Yacht Charters.com


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Christianity / Orientations / Heresy /
Sabellianism, or Modal Monarchianism as it also can be called, defined God as a single unity; that the Heavenly Father, the Resurrected Son (Christ) and the Holy Spirit were different modes or aspects of the one, single God, as if they were "faces" or "masks". Hence these three were not distinct persons or representations. God was a monad, an indivisible unity, and as Father he had expressed himself in Creation; God as the Son he had expressed himself in redemption of man; and God as the Holy Spirit he had expressed himself in sanctification.
In effect, Sabellianism claims that it was God himself who died on the cross. From this element in the doctrine, has another name been derived: Patripassianism.
Sabellianism points out that to God in the Bible, only the number One is ascribed. There is no mention of God being of the number Three.
Sabellianism was a rebellion against the Trinity, which often has been criticized as being a defection from Christian monotheism.
Sabellian doctrines would reoccur within Christianity up until modern times.
Information on Sabellianism is very uncertain, as the only surviving sources are from its opponents. Therefore, scholars judge the exact definitions of Sabellianism in a variety of ways.


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The NZ Herald reports that former National Party Member of Parliament Trevor Rogers has been jailed for failing to provide the court with design documents from a failed helicopter manufacturing company he used to own. The judge had this to say:
The evidence of Mr and Mrs Rogers that the (designs were) destroyed is not credible. Amongst other things, I do not believe that the most valuable asset of the company, and the asset that was most easily retained and hidden, would have been destroyed when large items were carefully hidden.
What I don't understand is how you can jail someone for something you suspect he hasn't done (i.e. provided documents that you believe exist) without any tangible evidence (since the court doesn't have the documents and so can't prove that they exist and therefore that he withheld them).
The guy may be totally guilty, but surely the onus is on the prosecution to actually prove that to some reasonable degree. I don't see how you can do that in a case like this. For the judge to simply say that he doesn't believe the defendant's evidence is credible strikes me as a claim that the defendant is responsible for proving his own innocence and a reliance on a very subjective standard of judgment. It doesn't make me feel very confident in the integrity of the system.


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Good Morning Images 2018, Best Good Morning Images For Whatsapp
Good Morning Images 2018, Best Good Morning Images For Whatsapp: If you want to wish a happy good morning to your partner, lover or friends then you have need some good morning images 2018 for them. You can wish those who are special for you good morning by the use of some latest and beautiful Good Morning Images 2018. Here we share nice collection of good morning images for facebook and whatsapp for our readers who want to wish their dear ones good morning.
"Awake, coffee, sunshine and another day off, aahhhhh, life is good!"
You can download these wonderful good morning images from our web page and then send or share them to your lover, partner, friends and family. You can also set these good morning images as your facebook or whatsapp profile pictures. We are share unique and best Good Morning Images 2018 with you.
So that you can send these Good Morning Images 2018 with your loved ones. We are also provided Good Morning Images, Good After noon Images, Good Night Images so that you can wish your loved ones whenever you want to wish them.
Good Morning Images For Whatsapp & Facebook
We are also share good morning, good night, good evening, good afternoon status and images with you. So that you can get each and every thing on our website and then you don't need to search here and there and your time is not wasted in searching.
You can send this best status or Good Morning Images 2018 to your friends, lover and family members by any messenger application. Now days Whatsapp, Hike is widely used messenger application of this era. So you can share Good Morning Images to your friend on this and wish them sweet good morning.
"Good morning A special cofee for you, mixed with love and affestion. have a beautiful day!"
Some people are habituate of sending these good morning images, wallpapers for Facebook or Whatsapp in early morning. It's a really good habit; now you can download good morning images for messenger application and send them to your friends. So here our team is going to share Best Good Morning Images for Facebook and Whatsapp and Good Morning Status. You can use these images to wish people in different ways. Hope you like these Good Morning Images for Facebook and Whatsapp. If you like our collection of good morning images then please share this on social media and with your friends. So share Good Morning Images to your friends and feel them that how especial they are for you.
Good Morning Images 2018:


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Our Proprietary Approach
Core Finance Team is led by a committed group of talented executives who have each gained over twenty years of experience working for the "Big Four" in the Healthcare industry. At Core Finance Team, we use our wealth of experience, personalized approach and dedication to our clients to deliver high impact results.
Our Core Values
Client Focused
Our firm was started so that we could better serve health care providers on a personalized basis. The same people selling the product will be leading the delivery of service to you. So if you have any questions or concerns, there will be no miscommunication or complex management structure to wade through, but rather a simple friendly phone call to make.
Our leadership team is comprised of members who have each successfully worked in the healthcare industry for over twenty years. This approach allows us to provide hospitals with valuable insights to their business, but also a broader perspective since each team leader has assisted over a hundred hospitals during his career.
Providing healthcare business advice is our passion. We have worked in the field and are dedicated to helping our clients achieve their goals. We enjoy the strong relationships we have with our clients and truly want to improve their operations and make sure that our results exceed our clients' expectations.
High-Impact Results
Our approach of utilizing a talented group of individual leaders devoted to our clients allows us to provide a service that adds more value than a large firm could. This sets our product apart and lets us work with a wide variety of hospitals and not just large accounts.
Meet Our Team
Our Services
Revenue and Regulatory Optimization
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Talk to us today about how to unleash your full profit potential and operational efficiency.


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Treasury's Lew to Europe
Let's hope this isn't a repeat of Woodrow Wilson at the Paris Peace Conference. In his first trip to Europe as Treasury secretary, Jacob Lew will try to persuade European finance ministers to focus more on growth and less on austerity. Europe's problems have not gotten better, with the sovereign debt crisis still roiling, unemployment rates rising, and the euro-zone economy shrinking.


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Roper Women's Brown Horseshoe Buckle Leather Belt
Item # 2589Y8
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- Roper Women's Brown Horseshoe Buckle Leather Belt
- Item # 2589Y8
Additional Information
Features & Description
- Genuine Leather
- 200 Brown
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- Sanding effect on surface of strap
- Alternating antique nickle finish domed nail heads and mini spots
- Non-stitched plain leather loop
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- 1 1/2"
Add some sparkle and flash to your outfit with this unique belt from Roper. This belt is made with genuine leather with embossed and burnished edges, has a sanding effect on the surface of the strap, alternating antique nickel finish domed nail heads and mini spots, non-stitched plain leather loop, and a horseshoe shaped buckle in silver finish. This eye-catching belt will add just the right amount of flash to your jeans. Add the perfect touch to your outfit with this belt from Roper.


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Would you like to unlock secrets like psychics and clairvoyants do? Would you like to have the answers to all your deepest, most important questions? Now you can harness your own psychic energy and get the knowledge that all fortune tellers have. You can do this by learning how to read tarot.
Tarot cards are a powerful form of divination that have been used by clairvoyants for centuries. They give the card reader great insight into someone's past, present or future. By learning how to read tarot, you can have this same remarkable insight. You will learn things that will amaze you.
To start learning how to read tarot, you should get a deck of tarot cards. You can buy them from a local psychic, magic shop, bookstore or on the internet. Find a deck you feel drawn to. This will help you make the most of your clairvoyant abilities.
Your cards will probably come with a booklet to help you learn how to read tarot. Read through this information and study the descriptions of the cards. You can get more detailed information in a tarot book. There are many books that explain the cards in more detail to help you understand their meanings and get to know how to read tarot.
Once you are ready to begin, choose a layout for your reading. There are a number of different card spreads, or layouts, that you can use. Whether you choose the cross formation, the horseshoe or a several rows will depend how the question you need an answer for. These are all good ways of how to read tarot.
To harness your psychic power and do well with your reading, clear your mind as you lay down the cards and focus only on your question. Let the energy flow through you. This concentration is important and will come more easily in time. It is an essential part of how to read tarot.
Another excellent way to learn how to read tarot is to speak with a tarot reader. These fortune tellers can share their own experiences with you and help guide your way.
Start your journey to wisdom now.


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As you may know, Anna Gilbert has a new album called "The Able Heart". You can buy it on itunes. Here's a teaser of her new album.
Anna Johnson did a cover of "Fix You" by Coldplay with her sister Julia for the 22nd Living Room Session.
Brooke Annibale had her song "You Don't Know" used in the episode 5 of the 5th season of Vampires Diaries.
Jess Penner will be releasing a new album in January called "Building Heaven". In the meantime, you can watch this teaser of her upcoming album.
Maria Zouroudis did a great cover of "Wrecking Ball" by Miley Cyrus. She brings it a further touch of sensitivity.
Meiko has a new funny official video for her song "Bad Things". That song was also used in the same episode of Vampire Diaries as for "You don't know" by Brooke Annibale.
ODyL has shared a new song on her youtube channel called "Les moins que bien". Her EP is available on itunes and her upcoming album should be out in 2014.
Roo & The Howl has released a new video for the song "To The River". This song has been one of my favorite by Roo & The Howl since it has been shared on soundcloud. I have listened to it over and over again. The melody is really beautiful. This great video reflects quite well the song.


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Creating coupons is very simple & effective. Helps build brand loyalty & helps retain your existing customers.
Follow these steps to create a coupon
-Log into your Orders2me Account
-Select Coupons from the dashboard
-Click Add Coupon
-Give your coupon a name & short description
-Choose if coupon will be flat dollar amount off or a % of the food order
-Choose if coupon applies free delivery or a free item once used on order
-Choose hours/dates for coupon to be effective
-Specify if there is a minimum order amount to use the coupon
-Decide on how many times a customer can use the same coupon code
-Decide on how many people can use this coupon code
-Specify if there is a maximum number of uses per day for the coupon
-Choose certain order type option of coupon code to be applied (takeout, delivery or both)
-Make sure to hit Submit to save coupon


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eG8 Summit in Paris Activists Fear Sarkozy's Efforts to Tame Web
French President Nicolas Sarkozy is holding a summit in Paris this week with some of the world's most powerful online luminaries. Netizens worry that Sarkozy's motives might be suspect -- and that he could leave a legislative mess like the one in Germany.
Nicolas Sarkozy is all too familiar with the pitfalls of the Internet. It wasn't long ago that the French president became the victim of an online attack himself, when unknown hackers hijacked his Facebook account and, in his name, circulated the false report that he would not be running for another term in office.
Sarkozy reacted with surprising equanimity to the hacker attack, poking fun at the many spelling errors in his adversaries' message.
But, since Monday, Sarkozy has been having a chance to discuss the attack in front of some very important people. He has invited three of the world's most powerful Internet luminaries to a forum in Paris: Eric Schmidt, the executive chairman of Google, the world's largest search engine; Mark Zuckerberg, the founder and head of Facebook, the world's largest social-networking site, with more than 650 million users; and Jeff Bezos, CEO of Amazon, the world's largest online retailer.
This is a rare gathering of such important figures. And the fact that Sarkozy, who is no stranger to the limelight, will actually be the one basking in the glow of great names is a particularly striking sign of the true importance of these online demigods. Indeed, people might not be able to point Paris out on the map, but they do know where the virtual realm of Facebook is: everywhere.
To make sure that the two-day G-8 summit starting Thursday in the coastal resort town of Deauville isn't just another routine meeting about the global policy agenda, Sarkozy chose to have this special "eG8" summit focused on the Internet before the main meeting.
The event, which is sponsored by major corporations and is being held under giant tents in Paris' Tuileries Garden, will see thousands of top industry executives and leading academics sharing their visions over two days. Anyone who was looking to hold the title of "co-chairman" of the event only had to fork over 250,000 ($350,000) for the privilege.
Sarkozy's War on the Web
Sarkozy has big plans. A joint resolution would essentially embrace the digital vision of Deauville, which the Elysée Palace refers to as a "thematic premier, that -- curiously enough -- has never been jointly addressed by the leaders of the industrialized nations."
The Frenchman has never shied away from theatrics and lofty words. And there's no denying that the major online companies -- which are mostly American -- and the world's top politicians -- and particularly the European ones -- either don't know much about or are suspicious of each other.
There are many topics to be discussed, including data protection, fighting terrorism, hacker attacks and, most recently, the seemingly manic desire of Apple and similar companies to hoard user location data.
Still, the Internet community views Sarkozy's effort to make "the Internet" the central topic of a G-8 summit with a great deal of suspicion. And perhaps with good reason: More than three years ago, Sarkozy declared war on the Web. At the time, he referred to it as a "Wild West" and characterized it as an "extralegal zone." In the style of an Internet Napoleon, he announced his intention to "civilize the Internet." Since then, he has pursued regulation with nothing short of missionary zeal.
Civilizing the Virtual 'Wild West'
In 2008, Frédéric Lefebvre, a member of parliament for the UMP, the conservative governing party, raged that, without strict controls, the Internet would remain a hotbed of "psychopaths, rapists, racists and thieves." After a recent meeting with Pope Benedict XVI, Sarkozy added: "Regulating the Internet to correct its excesses and abuses that come about in the total absence of rules -- this is a moral imperative!"
For some time, there has been a storm of new Internet-related laws and regulations designed to protect authors, copyright and internal security as well as to block certain websites. Sarkozy has aimed to "regulate," "cleanse" or at least "civilize" the same media sector that has often embarrassed him by publishing revealing quotes, photos and videos.
In 2010, France introduced a rigid three-step model for copyright violations and created a new regulatory agency called HADOPI. Under the legislation, anyone caught illegally copying music receives two warnings, the first by e-mail and the second by registered mail. At third violation brings anything ranging from monetary fines to an Internet ban. For the law's many critics, being blocked by one's Internet service provider is tantamount to a "digital death penalty."
The Sarkozy administration also turned its attention to unpopular content. A package of laws on internal security adopted in February includes the possibility of blocking certain websites, such as those displaying child pornography.
An Issue of Civilization or Control?
Sarkozy apparently wanted to leave nothing to chance for his big "Net G-8" appearance. Last fall, when then-Foreign Minister Bernard Kouchner was planning to host a conference on freedom of expression, Sarkozy sent him a letter warning him not to forget about his own goal of fostering a civilized Internet and reminding Kouchner that he, le président, had chosen this as a central issue of the France's G-8 presidency. As a result, the conference was promptly cancelled.
This track record has only fueled the current fears of online activists. The summit is "a farce," says Jérémie Zimmermann, the cofounder and spokesman of the advocacy group "La Quadrature du Net," noting that Sarkozy has made it clear that he wants to impose his authoritarian ideas about regulation on his G-8 colleagues.
La Quadrature du Net, together with other Net activists and initiatives, already called for protests in advance of the summit. Under the slogan "G-8 vs. Internet," the group argues that the world's governments are "uniting to control and censor the Internet."
It's always possible that Sarkozy will garner some support for his initiatives from his Internet celebrity guests in Paris. But, while in London last week, Google's Schmidt said that efforts to block domain names and Internet censorship could have devastating consequences for free speech. "I would be very, very careful about that stuff," Schmidt said.
- Part 1: Activists Fear Sarkozy's Efforts to Tame Web
- Part 2: German Missteps


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Brave boy!
By Mick Krever, CNN
A Palestinian-American teenager whose savage beating by Israeli police helped set off the current crisis described his shocking ordeal in an interview with CNN's Christiane Amanpour on Tuesday.
"They kicked me in the face. They kneed me in the face. They punched me in the face," Tariq Khdeir said. "They were beating me like they had no – they didn't know what they were doing. Why would they be beating me like that?"
Amateur video of Khdeir's beating went viral and caused widespread outrage; Israeli authorities say they have suspended one of the police officers involved in the case.
Khdeir was born and grew up in the United States; he was in the West Bank visiting family when three Israeli teenagers were kidnapped and found dead in the West Bank.
Khdeir's cousin, Mohammed Abu Khedair, was murdered in what was largely seen as a retaliatory attack…
View original post 473 more words


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- Kitchen & Home
From obsessive to people-pleasing, what does your coffee choice say about you?
Coffee - we all love it. Whether served steaming in a paper cup on the street or made at home in a coffee machine, us Britons knock back 70 million cups every day.
But the type of coffee inside those cups - from milky latte to short, sharp espresso - can say a lot about our personalities…
That's according to a study, which reckons - among others - that people who enjoy a milky latte are more likely to be people pleasers.
Some 1,000 people were surveyed by clinical psychologist Dr Ramani Durvasula. People were asked what type of coffee they drink and given multiple-choice answers to how they react to being in a queue, planning dinner parties and other things.
How about you? See yourself in any of the below?
Black coffee drinkers 'no-nonsense' folk
Ordering a coffee these days can be a complicated affair. Mocha or frappe, to syrup or not and hot or extra hot? The questions come thick and fast. So perhaps it's unsurprising to hear those ordering a simple, straight black coffee appreciate a simple life.
Yep, if you line up for an espresso on a Saturday morning the study has you down as a creature of habit who likes things to be straightforward. However, it also says you can be moody and abrupt - perhaps this is when the person in front of you is ordering a pimped-up, special-edition latte with more add-ons than a boy racer's Vauxhall Corsa.
Cappuccino lovers 'likely to be obsessive'
Always insist on ordering for your friends and choosing their syrup shot for them? You could well be ordering yourself a cappuccino, the research suggests. Boffins say if you like your coffee frothy then you're more likely to be obsessive and a bit controlling (ouch). Cappuccinos may be pretty mild - all milk and hot air - but as they say opposites attract… with more than the coffee getting in a froth according to this study.
Latte lovers 'like to please'
You may be hoping to have a latte when you meet up with your pal in town, but you could well end up letting them choose for you - as well as an outfit you really didn't like for that night on the town later. The study suggests latte drinkers are eager to please other people and happy to go out of their way to help others (was that you giving the young mum a lift down the steps with her pram while she held onto her chocolate shortbread?). With the study saying latte drinkers are more neurotic, you could well be worrying about whether you did the right thing regardless, whether you let your friend pick your coffee and clothes or chose your own.
Frappe drinkers like to 'make a statement'
Prefer your coffee blended with ice and drunk through a straw? Then you're likely to be a 'socially bold' type who likes to make a statement, researchers reckon. Your choices may not always be healthy, but you're always ahead of the curve and like the latest coffee to go with your latest fashions and furnishings. That's according to the survey anyway, which has frappe drinkers down as trendy Nathan Barley types with the latest trendy threads and gadgets - if only, I say. Anyway, next time I see a hipster in skin-tight jeans sauntering in under a mass of curly hair, perhaps I'll bet my mate a fiver that I know what drink he'll order…
The findings were published in You Are WHY You Eat: Change Your Food Attitude, Change Your Life.
See yourself in the above profiles or do you beg to differ? Let us know either way in the comments below….
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Related in Kitchen & Home


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Many hobbyists are frustrated with traditional swing-arm hydrometers that give inconsistent readings and only provide general guidelines for true salinity & specific gravity.
If you've experienced any of these typical concerns associated with hydrometers, it may be time to learn about the benefits of a C Scope Refractometer.
One of the most important parameters when keeping a marine tank is the salinity (salt concentrations) of your water. Natural sea water has a salinity of about 35 ppt (Parts per Thousand).
Monitoring the salt level should be accurate & precise to ensure a safe and optimal environment for tank inhabitants.
This simple integrated feature magnifies the scale over 2x compared to traditional refractometers for easy viewing.
Since water temperature effects salt readings, all C Scope Refractometers are equipped with ATC to give the correct reading even if your water temperature isn't.
The C Scope Refractometer will not only deliver the most accurate readings, it will do so consistently with the same test sample.
The ability to calibrate the refractometer allows you to cross reference your readings without ever have to second guess yourself.
The C Scope Refractometer was designed to provide a clear and sharp dual scale. This is accomplished with a deep color contrast between the blue and white to quickly distinguish your measurement.
The C Scope Refractometer is designed for testing the concentrations of saltwater by providing a direct
reading of the specific gravity and salinity in ppt.
It provides a reliable refractive index and salinity readings of total dissolved solids of aqueous solutions. The refractive index is the ratio of the the speed of light in a vacuum to the speed of light in the medium (saltwater) being measured. Light is slowed by passing through saltwater, so the index of the refraction is an indication of this speed change.
You simply add a few drops of your saltwater water over the prism and focus the eyepiece under direct lighting until you see a line that divides the scale from blue and white. This line indicates the specific gravity and the salinity. Clean off the prism and store. We recommend calibrating the refractometer before your next use. This tool will provide many years of TRUE readings, a must for all hobbyists!


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Here's an innovation story that follows a very familiar plot line, but the plot is so important that it provides an important reminder.
In an interview with a company's new CEO, a reporter tells us, the company has … "combined breathtaking engineering to create a whole bunch of amazing prototypes. But they rarely make it to market. That's because, over the last two decades, its culture has grown so competitive and insular, more consumed with getting and protecting an edge than pushing into riskier new businesses."
Does this sound familiar? It's the story of almost every successful business, where the balance between protecting the established core markets, and pioneering new ones, becomes a high stakes poker game played inside the company, with various camps competing for innovation, development, and marketing resources, and the established units almost always win.
The article goes on to say … "People were motivated to produce things they knew their managers would like, rather than take risks on new ideas that might fail. The company's money-minting core offerings sucked up talent and attention while newer ideas got overlooked."
But what we all know is that the established units win for a while, until major shifts in the market leave them flat-footed. In fact, this situation is so common, the reporter goes on to note that "It has become accepted wisdom in Silicon Valley that large, successful tech companies can't reinvent themselves. Many have attempted to engineer comebacks, and the industry is teeming with failed empires that have evolved into middling businesses on the decline."
Is this a story about HP? Or Blackberry? Or Yahoo? It could be any of those.
Actually, though, this article is about Microsoft. It's a great piece in the latest Wired, a profile on the new CEO Satya Nadella. The article is quite sympathetic to Nadella's task, and frames him as a smart and sensitive executive who could well succeed in the monumental task that he is confronted with.
Nadella himself describes three layers of critical organizational performance, Concepts, Capabilities, and Culture, and notes that while Microsoft is awash in great concepts, "You need a culture that is fundamentally not opposed to new concepts and new capabilities."
While "Not fundamentally opposed" puts the bar pretty low, he has a good point, and it certainly frames the work that he wil need to be doing in the coming weeks and months.
The next article that I'd like to read is about the how. That is, How does Nardella expect to achieve this in the Redmond behemoth? The Wired article unfortunately doesn't say much about that. But we do. This is squarely in the InnovationLabs competence area, and we've written about it extensively, including in the two books mentioned to the right, Agile Innovation and The Innovation Master Plan.
Among the case studies we cover in Agile are Wells Fargo, Nike, and NASA's Apollo Program, each tackling a different variation on the same situation, and each providing new insights.
At Wells Fargo, it's about how to take risks in a highly regulated and controlled banking environment that is nevertheless subject to monumental forces of change; Steve Ellis gives us the metaphor of snowboarding to explain how he makes the organziation move faster.
At Nike, it's about pushing a massive volume of new products out into the market, and then listening carefully to find out where there is resonance, and shifting even the brand identity when the company and the market become out of sync.
And for Apollo, it's about how thousands of engineers and scientists created something of monumental scale, something that had absolutely never been done before – the moon landings – while balancing innovativeness and control.
Nadella is facing a task that has elements of all of these stories, and we will be fascinated to witness the new chapters as time, history, and the market write them.
PHOTO: Buzz Aldrin on the moon. Photo by Neil Armstrong. Courtesy of NASA.
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http://www.w3.org/Bugs/Public/show_bug.cgi?id=14985 Summary: Specify how autoincrement + empty keypath works Product: WebAppsWG Version: unspecified Platform: PC OS/Version: All Status: NEW Severity: normal Priority: P2 Component: Indexed Database API AssignedTo: email@example.com ReportedBy: firstname.lastname@example.com QAContact: email@example.com CC: firstname.lastname@example.com, firstname.lastname@example.com What should happen for the following calls? store = db.createObjectStore("mystore", { keyPath: "", autoIncrement: true }); store.add(1); // store an item with key 1 store.add("foo"); // store an item with key "foo" store.add({foo:"bar"}); // throw It seems like if you have a objectStore with an empty keyPath and autoIncrement set to true, you'll either a valid key as the raw value, in which case that value is used as key. Or you'll specify an invalid key as value, in which case we should throw since it's not a valid key. In neither case would we actually use the key generator. It seems to me that the simplest solution is to not allow an empty keyPath and autoIncrement set to true. We would of course still allow no keypath and autoIncrement set to true. I.e. we'd still allow out-of-line keys together with autoIncrement. As well as non-empty keypaths with autoincrement set to true. -- Configure bugmail: http://www.w3.org/Bugs/Public/userprefs.cgi?tab=email ------- You are receiving this mail because: ------- You are on the CC list for the bug.Received on Tuesday, 29 November 2011 00:23:37 UTC
This archive was generated by hypermail 2.3.1 : Friday, 27 October 2017 07:26:37 UTC


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Cruise Family – We're in Heaven 12″
Cosmic synth music requires regular transfusions of reverse strategies, weird ideas, and bad blood to stay mutating and alive. And while there's no shortage of projects engaged in this pursuit, the life/art arc of Austria's Cruise Family cleaves closest to the slipstream we identify with. His early tapes were bubbling fountains of faded trance textures, like a rave reimagining of Monopoly Child Star Searchers' celestial cassette mysticisms, but We're In Heaven extrapolates that DNA into more dynamic forms. The sidelong A piece, "We're In Heaven," coasts on a carpet of lush galactic synthesizer bliss and metronomic 909 pulses, like someone cross-mixing the best of Software's Electronic Universe over a slowed-down Model 500 instrumental. The B is split into two tracks: the paranoid warehouse techno skeleton, "Be Part Of It," and "Gone By Dawn," which sequences floating new age fractals over a hollow acid handclap, like the spectral shell of some classic sunrise anthem. The fidelity and craft of these cuts are distinct from anything else in the Cruise Family tree, and the best embodiment of his unique matrix of influences. Black vinyl 12 inches in jackets with laser humanoid artwork by Stefan Kushima.


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REMINDER: 2nd annual 'Burien UFO Festival' is just one week away!
REMINDER: You've got just one full week to get your homemade UFO built, and your Man/Woman in Black – or Alien – costume ready as the second annual 'Burien UFO Festival' is coming to Olde Burien next Tuesday night, April 1, starting at 7 p.m.!
Like last year's wildly successful inaugural event, this night will include a 3-D flying saucer, awesome Flash Mob dance, cash costume contests for Best Man in Black, Best Woman in Black and Best Alien, live music, and much more.
Organizer John White will be giving out $100 cash prizes to the following "Bests" in a costume contest for:
- Best Man in Black
- Best Woman in Black
- Best Alien
"It's going to be another awesomely humorous yet strange night with great live music, another Flash Mob dance, costume contest and high weirdness," White told The B-Town Blog over an encrypted, NSA-proof phone line.
Here's what to expect:
- Best Alien/Man in Black/Women in Black costume contest
- Bring Your Own UFO contest
- Flash Mob Dance
- Live music from the Pink Torpedoes
- Photo shoot of all Men/Women in Black & Aliens at Tin Theater, starting at 9 p.m.
- Help "The Maury Island Incident" raise money for film festivals/marketing by buying customized t-shirts – just $20 each!
Be sure to "Like" the Burien UFO Festival Facebook page here: https://www.facebook.com/BurienUFOFestival


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kenya, nairobi, slum, poverty, violence, trash, poor, church, virgin mary, legion mariaAdd to Lightbox Download
Members of the Legion Maria Church pray. Mathare, one of the poorest slums in Nairobi. Running water and electricity are scarce and trash and human waste fills the streets. Many people have no jobs and those who do work can earn less than one dollar a day.
- Filename
- Rochkind_KenyaMathare31.JPG
- Copyright
- David Rochkind
- Image Size
- 4368x2912 / 1.8MB
- Contained in galleries
- Mathare Slum / Kenya


text: Canada's Walk of Fame is excited to announce that Ayaz Virani of North York, Ontario is the Grand Prize winner of the 2016 Emerging Artists Music Mentorship Program, supported by RBC. The program, valued at more than $100,000, provides musicians with the opportunity to grow and develop their skills and industry connections, and take their budding music careers to the next level. Seven talented finalists were selected from more than 350 hopefuls from coast-to-coast who submitted applications that ranged across all genres of music.


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Reserve Bank of New Zealand Banknote
Pick-185. Dated 14. Prefix BK.
Reserve Bank of New Zealand $5.00 Polymer Banknote. Issued 2014. Signature G. Wheeler. Governor (2012 - Present).
Water Mark Queen Elizabeth II. Printer: Note Printing Australia.
Your browser does not support inline frames or is currently configured not to display inline frames.


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New Canon Printer Is Simple To Use
Canon's new MP490 printer is cheap and "simple-to-use", a product reviewer has claimed.
Writing for PC Advisor.co.uk, Susan Silvius said that the printer should appeal to both small-to-medium enterprises and home users.
Features singled out for praise in the review include an LCD screen which shows an animation illustrating how ink cartridges can be changed.
The small screen also has a "compass-style" navigation wheel with a number of well-labelled buttons to enhance the user experience.
Ms Silvius said: "The £79 price range of the Canon Pixma MP490 MFP means tradeoffs. The high ink costs are bearable if you print infrequently; otherwise, look for a model with lower-cost inks (and a higher purchase price), such as HP's Officejet 6500 Wireless.
However, the expert criticised the cost of ink cartridges for the machine and suggested that if people are looking to print regularly, then a different device could be preferable.
The printer does not have Ethernet or automatic duplexing.
Features include Copy Facility Present and Flatbed CIS Colour Scanner.


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DRDO CEPTAM 08 Hall Ticket 2016, www.drdo.gov.in, DRDO CEPTAM Admit Card, DRDO CEPTAM 08 Admit Card 2016, DRDO CEPTAM Hall Ticket 2016
Defense Research and Development Organization (DRDO) has recently decided to release out the admit cards for the CEPTAM exam. Defense Research and Development Organization (DRDO) is an agency which comes under the agency of Republic of India. This agency was fully set up in the year 1958 by the Technical Development Establishment along with the Directorate of Technical Development and Production. DRDO agency has their main headquarters situated in New Delhi.
डीआरडीओ सीईपीटीएएम (DRDO CEPTAM 08) परीक्षा का परीक्षा पत्र Admit Card Download करने के लेय उपलब्ध है। आप DRDO की Main वैबसाइट से इसे डौन्लोड कर सकते है।
In the past few weeks the agency released online application forms for the unoccupied seats. The seats vacant in the agency are for the posts of Senior Technical Assistant, Technician A and Admin & Allied. For Senior Technical Assistant there are 564 seats vacant, for Technician A there are 345 seats available and for Admin & allied there are 233 vacancies available. All these vacant seats make a total of 1142 vacancies available in the agency. For appointing the eligible contenders in these seats, a written exam will be conducted.
DRDO CEPTAM 08 Hall Ticket 2016
The date for the CEPTAM written examination has not mentioned till now but the agency has finalized that the exam is going to be conducted in few days. The DRDO written exam will be only organized in whole country as the agency comes under the central government of India. Examination centers will be allotted within various states & districts by DRDO in the country only as per the candidates' preferences and choices. In the written exam, objective questions will be asked as per the subjects. 3 hours' time period will be allotted to the participants to complete the exam.
But to appear in the exam, the participant must have their respective DRDO CEPTAM 08 Hall Ticket 2016. When the applying process will over, contenders were more eager to get the admit cards. They were trying find out the admit cards on the different websites by surfing their different terms. But to make this process much simpler, the agency is going to release the admit cards on the authorized link of the Defense Research & the Development Organization (DRDO). Candidates must visit the site and download the respective hall ticket/admit cards, because the candidates who will be without the admit cards will not be given permission to sit in the examination hall.
DRDO CEPTAM Admit Card 2016
Admit card or we can say Hall Ticket is one of the neediest document for every exam & to every single participant. Admit cards/Hall Ticket contains lots of important information about the exam and the appearing candidates. Details like name of the examination center, date for the conduction of exam, full name of the participant, exam venue, & the identification of the candidates are printed on the DRDO CEPTAM 8 Hall Ticket 2016. Some required instructions are also shown on the hall tickets (Admit Cards), which the candidates need to follow in the examination hall. It is required by every single candidate and for all participants it will be shown on the official link page of the organization. Candidates must get the admit card from the agency's main website. As the participants who won't be having the admit cards will not be permitted to take part in the exam.
DRDO CEPTAM Hall Ticket 2016
Firstly, www.drdo.gov.in visit this official link of board.
And in some seconds the home page will be appeared.
On the main page look for the most appropriate link for the DRDO CEPTAM 08 Hall Ticket 2016.
After clicking on the link, select the respective DRDO CEPTAM Hall ticket .
After entering all such details on DRDO Website click on submit.
Within fraction of seconds the respective admit card for the entered details will be visible on the screen.
Candidates can download it by just clicking on the download link.
After the complete download, take a print of the CEPTAM Hall Ticket for further use.


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First, our schedule for the week (3/21 - 3/25)
Lunch (11AM - 2PM); see below
Dinner (4PM - 8PM) - Warren & Greenwich St (Tribeca)
Mon 52nd St between 6th & 7th ave (Midtown West)
Tues 55th St between 3rd & Lexington Ave (Midtown East)
Wed Hudson & King Street (SoHo)
Thu Amsterdam Ave & 116th Street (Uptown)
Fri Water & Broad (FiDi)
What happened to Spring? We wanted to add our spring addition to the menu, it will just have to wait :)
The Burger Of The Week is the MARITAL BLISS - Seasoned Grassfed Angus Patty with Philadelphia Cream Cheese, Jalapeno Peppers and Applewood-smoked Double Thick Bacon.
On Saint Patrick's Day, we brought back the THE BOURBON BURGER - Marinated Kobe Patty and marinated onion jam and oven roasted tomatoes! Once again, the people loved it. So.....we will bring it back for another day. It will be available this Friday :).
We will also have our delicious Homemade Dessert Treats and Soups!
As always, if the weather is not great & you would like to shorten the wait time, you can always place your order 10-15 minutes prior to pick up. Place your Pre-order at 917.292.9226
Email (firstname.lastname@example.com) or call us to have us cater your private or corporate event.
Don't forget to follow us on twitter.com/fritesnmeats and facebook.com/fritesnmeats peeps!
Don't forget to log on to our site, fritesnmeats.com, via your smartphone as it is now powered by MobileMeteor!
From there, a map will appear with our current location.
You can also clearly view our blog for schedule and menu and rate your experience with us!
We look forward to hearing what you think.
Have a great week everyone & lets do the Spring Dance!
Sunday, March 20, 2011
Did they say spring is here?
Posted by fritesnmeats at 9:33 PM
Labels: burger, Events, Festival, Food, fries, mobile devices, new york city, truck
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|By Admin1 (admin) (ppp-70-245-110-11.dsl.okcyok.swbell.net - 188.8.131.52) on Friday, February 10, 2006 - 4:46 am: Edit Post
1981: Cheryl Jovel served in Sierra Leone in Freetown beginning in 1981
Returned Peace Corps Volunteer Cheryl Jovel can be contacted at Blindbizaaoldcom
Country of Service: Sierra Leone
Training Group: I don't know!!
Cities you served in: Freetown
Arrival Year: 1981
Departure Year: 1983
Work Description: Teacher for Blind Children and Adults
Bring us up to date on your life after the peace corps:
I worked for 2 years in Bolivia as a teacher for the Blind and then got a
masters degree in International Administration from S.I.T in Brattleboro,
Vt. I did an intership with the United Way in Jamaica for 6 months then
joined the Caribbean Council for the Blind for 2 years as an instructor for
new teachers of the blind throughout the Caribbean. I got married in 1989
and have 2 kids. I currently am the office manager for my husband's
Family Practice office in St. Pete, FL
Any thoughts you have now looking back on peace corps days?:
I griped a lot about it while I was there but now I am so glad I did it....my
whole life changed for the better.
Anyone you are looking for or would like to hear from?:
Anyone who served in Sierra Leone
Any message for returned volunteers?:
Look me up!
Originally posted: January 31, 2002
Looking for another volunteer who served with this Group?
If you know this volunteer or someone from his or her group and are looking to reconnect with them,
leave a message above where it says "Create New Conversation."
If this is your profile, post any additions, updates or corrections to your profile below where it says "Add a Message."
RPCVs: To add your own RPCV profile click here.
| The Peace Corps Library
Peace Corps Online is proud to announce that the Peace Corps Library is now available online. With over 30,000 index entries in 500 categories, this is the largest collection of Peace Corps related stories in the world. From Acting to Zucchini, you can use the Main Index to find hundreds of stories about what RPCVs with your same interests or from your Country of Service are doing today.


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About Cameron's
Setting the standard
Cameron's Catering burst onto the Champaign scene in October 1994. Ever since, Cameron's has been guaranteeing the best possible food and service as one of Central Illinois' largest privately owned and operated caterers. Keith Cameron Smith (owner and founder) alone brings more than 31 years of hospitality experience to the table, including more than 20 years in the catering industry.
No detail is too small
With this homespun philosophy as our guide, Cameron's takes great care and interest in perfecting every aspect of your event—as though it were our own. And we bring that passion to work with us every day to ensure that each event—from the casual to the extravagant—is deliciously prepared, beautifully presented, and worry free for you and your guests.
We only use cookie cutters for some of our desserts
Just as no two brides are alike, neither are any two wedding receptions. Cameron's designs your event's menu, decor, and presentation specifically for you and your guests. The example menus on our site are only the beginning—think of them as a jumping off point for customizing to your own unique style and flavor. If you are ready for the five-star treatment, please contact one of our event professionals for a free consultation today.


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Galapagos Passion Explorer and their experts of high quality service
When cruising on Galapagos luxury charter, you will receive an experience like no other in this exotic area of the world. We understand the type of holiday that our demanding clientele wants and deserves. Every member of our dedicated staff is here to provide you what you want. Unlike other island cruises, every staff member on the Galapagos Passion Explorer private charter is fully bilingual to give you the best quality service. Our bilingual professionals come from diverse fields to ensure your complete satisfaction. On board staff usually consists of a crew manager, hosts and hostesses, gourmet chef, photographer, naturalists and local guides and dive master instructor.
It's all in the details and the logistics when on board a Galapagos Passion Explorer luxury yacht. Established by two entrepreneurs who understand the need for reliable information and complete service at this unique destination,Galapagos Luxury Charters staff works long before your vessel ever leaves port to make sure that every detail and request will be met.


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Two Members of Legislative Assembly from the Telangana Rashtra Samiti on Friday climbed the Legislature Party building in the Andhra Pradesh assembly premises and threatened to jump off if a separate state of Telangana was not created immediately.
Other MLAs of the party burnt an effigy of the 'Seema-Andhra government' near the legislators' entrance gate in the assembly premises.
Holding black flags, the two MLAs -- K Sammaiah and D Vinay Bhaskar -- climbed the terrace of the building and also demanded that all the Telanagna protagonists arrested by police in connection with the Chalo Assembly programme on Friday be released immediately.
Assembly marshals brought the two MLAs down after an hour.
The AP assembly was, meanwhile, adjourned for the day without transacting any real business as TRS, Telugu Desam Party (Telangana Forum), Communist Party of India and Bharatiya Janata Party legislators disrupted the proceedings on the statehood issue.
Two adjournments did not restore order and when the House re-assembled the third time, Speaker Nadendla Manohar asked Finance Minister Anam Ramanarayana Reddy to move two Bills.
After the formality was completed amid the din, the House was adjourned for the day.
Nearly 20 TRS, BJP and CPI MLAs, who were sitting at the two gates of the assembly after it was adjourned, were detained.
Earlier, the Telangana issue stalled the proceedings of the assembly, which resumed its budget session on Monday, even as Hyderabad city remained under beefed-up security in view of the Chalo Assembly call given by pro-Telangana activists.
The assembly proceedings were adjourned twice on Friday morning, as TRS and TDP MLAs from the region stormed the Speaker's podium, demanding the immediate creation of a separate state.
Sporting black T-shirts, TRS MLAs were up on their feet as soon as the day's proceedings began and demanded that Parliament immediately pass a bill for the creation of Telangana state.
The TDP legislators from the region joined the issue, forcing the Speaker to adjourn the House for 30 minutes.
The BJP and the CPI joined the TRS and the TDP members in the Well of House when it re-assembled after an hour.
With no sign of order even after he made repeated appeals to the protesting members, the Speaker adjourned the House for another half an hour.
The treasury benches wore a deserted look as most members, including Chief Minister N Kiran Kumar Reddy, Deputy Chief Minister Damodar Rajanarasimha and ministers, did not enter the House.
Only three ministers could be found inside the House as the Opposition din continued to paralyse the proceedings.
Image: MLAs protest at the Andhra Pradesh assembly premises | Photograph: SnapsIndia


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Hello everyone :)
Thank-you for the lovely comments last week :)
I'm back today with my C4E DT card. This weeks challenge is called
We would like to see your sweet themed projects, anything with sweets, chocolate or cake on.
Our sponsor Crafty Sentiments Designs has kindly donated a choice of 3 digital stamps to one randomly selected winner.
For my card this week I have used a C4E digital image called A Balanced Diet.
The lovely Nella is balancing some yummy cupcakes which are topped with gems!
I have coloured Nella with Distress ink and just a hint of stickles ;)
The DP's are Bo Bunny and I have used a Bo Bunny sentiment stamp along with a My Minds Eye Butterfly stamp and Martha Stewart punched butterflies.
A few flatback gems and generous helpings of Stickles complete the card.
Hop over
to join us and see what the fabulous DT girls have done this week.
Thanks for stopping by have a great weekend!
Michele x


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ObituariesGilbert (Gil) Chafe
Passed suddenly away surrounded by his loving family at St. Clare's Mercy Hospital on Friday, March 6, Gil Chafe, a retired employee of Memorial.
Robert H. (Bob) Morrow
Robert H. (Bob) Morrow passed away at his home in Grand Falls-Windsor on Feb. 21 at the age of 81. Bob retired from his own business, Morrow Nurseries, in 2000. He started the business in 1949. From 1977 to 1983, Mr. Morrow served two terms on Memorial's Board of Regents.
The Hon. Justice Pleman Lloyd Soper
Judge Lloyd Soper passed away in St. John's on March 6. Dr. Soper graduated from the former Memorial College (Parade Street campus) in 1940. He received an honorary doctor of laws degree at the special convocation for the official opening of the School of Fine Arts, Sir Wilfred Grenfell College, in October 1988, and was named Alumnus of the Year in 1995. Dr. Soper was a familiar face on the Corner Brook campus. His most public contribution to Sir Wilfred Grenfell College was the establishment of the Honourable P. Lloyd and Elizabeth Soper Scholarship in 1994.


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A team of scientists have finally filmed a sight they have been waiting to see for decades: a drop of extremely slow-moving pitch break off and fall.
The team running the pitch-drop project at Trinity College in Dublin witnessed it detach at roughly 5 p.m. on July 11, according to Nature News.
It may not seem like a hugely momentous event, but moments like these only come around once a decade or so, and the drop has never been filmed before. Paying close attention to this slower-than-molasses experiment gives scientists an extraordinary glimpse into the physical properties of an ordinary substance.
"We were all so excited," project leader Shane Bergin told Nature News. "It's been such a great talking point, with colleagues eager to investigate the mechanics of the break, and the viscosity of the pitch."
Pitch, also known as asphalt or bitumen, moves so slowly at room temperature that it appears to be a solid. Watching the drop allowed the team to estimate the viscosity of the pitch: It flows about 20 million times slower than honey and more than 2 billion times slower than water.
They think that the funnel and pitch was set up in 1944 by Nobel laureate Ernest Walton who was a professor at the school. They think he wanted to use the experiment as an educational tool.
When first set up it took three years for the blob of pitch to begin flowing out of the funnel, and then another seven to ten to form a drop. The set-up lay neglected for years, until scientists literally blew the dust off the pitch and began watching it again.
The flowing pitch is one of the world's longest-running scientific experiments. Last April, they set up a Web cam so people around the world could watch the drop finally break off and fall.
While it has taken years to flow out of the funnel, it only takes a tenth of a second for the drop to dislocate and fall. Seeing it happen is so rare that in the 83 years that an even older pitch experiment has been running at the University of Queensland in Brisbane, Australia, no one has witnessed any of the nine drops that have fallen.
Watch the Trinity College drop fall, the first time this historic event has ever been captured on camera:
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Business Insider: Science


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In a mailing from the McCain Campaign, Senator Joe Lieberman detailed his support for Senator McCain...
"Time and time again John McCain has put his country first. He refused early release when he was held captive in Vietnam. He continued to put his country first as a national leader in the U.S. Senate. He put country before party when he fought to pass campaign finance reform, sought a bi-partisan solution to the immigration problem, and consistently supported pro-environment policies. His courage to stand up to the failed Iraq war plan of Secretary of Defense Donald Rumsfeld and lead the fight for a new strategy in Iraq will go down in history, and it saved American lives. These were not always the easy things to do. In fact, they were usually very difficult, and often threatened his political career. But John McCain did what was right."
Senator Lieberman is chairing a new organization called Citizens First for Senator McCain's campaign made up of Republicans, Independents and Democrats who support a McCain presidency.
Friday, June 6, 2008
Lieberman Explains McCain's Appeal to Independents
Posted by kmorrison at 8:28 AM
Labels: citizens first, country first, democrats, joe lieberman, mcain lieberman, mccain independents, moderates, rumsfeld, vietnam
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WECBOF is a voice and a resourceful network for entrepreneurs, corporates, and government to collaborate in an effort to build a stronger and more prosperous Western Cape economy. In 2013, the WECBOF board decided to re-focus and re-design the structure of the business to ensure that it remains relevant, innovative, and progressive.
Currently, with so many organisations similar to WECBOF in the market, it makes sense to collaborate rather than work in isolation to serve a broader business community.
The premise of WECBOF's new Organisational-Operational Structure 2013 – 2020 is one of lateral interaction, i.e. working collaboratively with other organisations to achieve a common objective. A modern approach to management and strategic approach and implementation has been adopted by WECBOF which is visionary, resourceful, and inclusive of all stakeholders for the advancement of the local economy.
With the launch of the new BEE Codes in May 2015, government has made a strong statement about its commitment to growing and developing entrepreneurs and small business. WECBOF wishes to align itself with the requirements of these codes to ensure that it can advise its members adequately as to how these codes have to be implemented and monitored in the working environment.
Join us on this new and exciting journey; together we can achieve so much more…
HistoryAccess Token not set. You can generate Access Tokens for your Page or Profile on fb.srizon.com. After generating the access token, insert it on the backend
Hope – the one word which summed up the feeling in the hearts of millions of disadvantaged people of South Africa on 27 April 1994.
The results of our country's first democratic election sowed the seed for great expectations, but would legislative changes be enough to bear fruit for all in the longer term? It became apparent that Black business leaders needed to play a more meaningful role in addressing the imbalances of the past by consolidating their efforts to nurture and develop economic growth.
It very quickly became clear that there were great imbalances in South Africa's economy. It was out of this imbalance and wealth disparities that WECBOF was founded. Questions were raised around the relevance of the organisation, given that there was already a number of Chambers of Commerce and formalised,successfully operating businesses, in the marketplace.
However, WECBOF fulfilled business requirements such as access to finance, training, and opportunities, which became its unique selling propositions at the time.
WECBOF was formed in January 1995 by local businesspeople, academics, and other prominent members of the local business community. Its formation was the result of months of planning and discussion around how previously disadvantaged small-and medium-enterprises could be meaningfully integrated into the economic landscape of the Western Cape Province, and operate as successful and sustainable enterprises.
WECBOF's mission was established with its primary objective being to effectively contribute to the economic and social wellbeing of the province.


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Welcome to the Vinci Hotel Restaurant.
Air-conditioned and elegant interior, welcoming and professional staff along with a great variety of delicious national and international dishes provide a wonderful┬Ā setting for business meetings,as well as social and family gatherings.
We organise banquets, wedding receptions, ceremonial dinners, business lunches and other events.
Opening hours: 1 pm ŌĆō 10 pm.