text:
Rated 3.9 out of 5 by 739 reviewers.
Rated 5 out of 5 by Danna00 My favorirty Starwood Property in NOLA The Sheraton is by far my favority Starwood property in the city. The staff is always friendly, professional and goes out of their way to meet my needs. I love the Club Lounge and Fitness Center/pool. The rooms are spotless and well appointed for the business traveler. Brandon, one of the managers, was especially helpful during my last stay and went out of his way to ensure that my boyfriend and I had a great experience. August 19, 2013
Rated 4 out of 5 by Brian1962 Great location, comfortable rooms Love this hotel's location! July 14, 2014
Rated 2 out of 5 by None27 Disappointing I don't usually write negative reviews but I am really disappointed in this hotel. I usually stay at Sheraton's most of my travels. Here is a situation that should summarize the attitude of the staff in this hotel. Every time I would come back from my conference, I would find water on the floor of the bathroom. I thought I didn't close the shower curtains properly and didn't think much of it. Last day, I realized that water was coming from the ceiling every time the occupant of the room above mine would use their bathroom. Here is the kicker: Upon telling the staff, I was told that a call to maintenance would be placed. No excuses. No sorry for the inconvenience. Nothing. Weird to say the least. Cleanliness of the bathroom was also below average. Isolation between the rooms was abysmal. I could hear the guy next door coughing!!! I will chose a different hotel when in NOLA. January 29, 2014
Rated 4 out of 5 by Glenn1503 High Recommendations Over all the facility lived up to the 4 * rating you have. Things that made me not want to give 5*. No ice machine and not told apon check in on 15th floor. Apon entry of room and general review of room under beds needed to be vaccumed (badly). On the second day there I wanted to make coffee and old coffee flter still in machine from last guest still there.. My first time to have breakfast..there is more to say.... September 3, 2013
Rated 4 out of 5 by catperson2 Dont' get room service Had room service for breakfast. Prices were outrageous. My spouse's eggs and toast were cold. Coffee was lukewarm I ordered yogurt and fruit .....got a small container of Yoplait strawberry yogurt. June 29, 2014
Rated 5 out of 5 by Sandy49 This hotel is wonderful! The hotel room/bed was very comfortable. Close to all attractions/restaurants in French Quarter. The staff was very helpful and friendly. July 4, 2014
Rated 5 out of 5 by Toni28 This hotel was perfect for the conference The layout and location of the hotel worked best for this conference. July 9, 2014
Rated 4 out of 5 by Obi25 NOLA EXPERIENCE The Sheraton New Orleans was a great place to stay. Walking distance to everything in the area. The staff was excellent and very nice. My tub was stopped up in one room and they moved me to another room no problem. I will definitely be staying there when I return. June 16, 2014
Rated 5 out of 5 by Harleychick1 Essence 2014 "GREAT"!!!! Hotel was in a great location and the service was excellent!!!! July 8, 2014
Rated 2 out of 5 by JackRyan Did not get value for money, disappointed Toilet was not cleaned properly when I first walked into the room, had to flush it off myself. Trash box was in the hallway, right by the main door and I would have tripped. Most times, during both day and night, I had to wait for a long time to be seen by someone at the check-in desk and even the concierge. I don't think I will stay here again, pretty disappointed. April 30, 2014
Rated 2 out of 5 by JGZ2 Marginal review The electrical failure which closed down the upper level elevators for a day and a half and resulted in no electricity and air conditioning for a number of hours on the second day put quite a damper on out stay at the hotel. June 22, 2014
Rated 2 out of 5 by MikeV123 UnHappy Most likely our last time at this hotel. Was charged for Club $20.00 per day as other were just opening the door and letting people in. As a gold member I think we should not have to pay for that service. Also room next to elevator and ice machine. My profile says please not next to elev or ice machine. never offered upgrade or anything at check in. was charged an additional $50 dollars for what I don't know. It's on my AmEx statement not happy at all with stay. ect....... I guess the Hotel was hoping I would not review my statement Tried to talk with someone at the hotel and was put on hold then the never called back. Went down to front desk 3 people checking in and out with lines of people. Just not what I expect. !!!! Parking $42.56 outrageous took 45 min to get our car. I could go on but will save it for the SPG people. Next stay in New Orleans will not be at this hotel.!!!!!! August 19, 2013
Rated 4 out of 5 by chrism28 This hotel is good Our stay was find. The room were nice, clean and the bed was comfortable. The pool was small. Be alert in the lobby a person came up to me and asked me for money. June 29, 2014
Rated 5 out of 5 by Lightsout7 Great view This property has a great view of the city and the river. July 7, 2014
Rated 4 out of 5 by NHQ54 Great location Excellent amenities, located close to the French Quarter and the River Walk. Staff are friendly and helpful, I would definitely stay here again. June 30, 2014
Rated 4 out of 5 by tc14 great location, comfortable and clean The hotel was clean and the staff excellent. The room was comfortable and relatively quiet. One disappointment - the main restaurant in the hotel closes at 2pm and not open for dinner. With bad weather outside and a need for a quiet evening, this was a problem during the stay. June 8, 2014
Rated 5 out of 5 by hebrews11 Great features and location I really enjoyed my stay at your hotel. This was my first visit to New Orleans and the comfort and service of your hotel made my visit extraordinary! I was well within walking distance of many of the tourist attractions. The French Quarters were everything I had been told to expect. Thanks for making this experience such a great one! May 16, 2014
Rated 4 out of 5 by Brandy72653 Nice Hotel The hotel is very nice and comfortable. I would recommend this to anyone staying in New Orleans. Staff was very friendly and accommodating. The fire alarm did go off during our stay (only in our room) but staff promptly took care of the issue. The worst part of the say was in the late evenings. The amount of homeless people in the street off Canal, was difficult to see. April 10, 2014
Rated 3 out of 5 by JacobP Fine hotel, conveniently located The Sheraton New Orleans is a fine hotel, centrally located in downtown New Orleans. It's a big hotel, and New Orleans almost always has something going on during the weekends--a convention, or a festival, or whatever. That can cause the hotel to get really crowded, and feel overwhelming in the lobby. Other than that, it's a fine base for exploring an incredible city. April 14, 2014
Rated 2 out of 5 by NYJC Disappointed - I expected more from a Sheraton The hotel is generally acceptable, but I was quite disappointed in the cleanliness of the room. It looked like it had not been dusted in months and there was a half-eaten granola bar under the bed. The room was only half-cleaned during my stay and the shower mat, ice bucket, etc. were not changed. The previous guest set the alarm, which was left on and not cleared by the staff (would have been nice!). The toilet ran non-stop (which I did report and was called in to maintenance.) The elevators were pretty high-tec, but were very slow. The pool is small and inconvenient to get to and the hot tub was closed. I had a few banquet style meals during my stay and I was very disappointed at the quality of the cooked food. I realize it is hard to serve a hundred or so people, but the chicken was almost inedible. On the plus side, the staff was great and the location is good. The room is fairly modern and comfortable. March 24, 2014
Rated 5 out of 5 by 52Nancy This hotel is a "green" For those of you who are "green", this hotel offers "green" options and if you are an SPG member, you get rewarded!! Upon checking in, we were given the option of maid service every day (new sheets, towels, products, etc.) or going "green" We chose to go "green" and received 1,000 SPG points for doing so along with a $5 hotel voucher for each day. We could always get extra towels, coffee, etc if we needed them, which we didn't. We gave up new sheets and thus saved on water, detergent, electric, gas, etc. which is great for the planet, and we were then rewarded for doing it!! Great idea SPG!! Besides this, the hotel is in a great location, has a huge, bright, Lobby bar, hotel computers have great connectivity, there's a Starbucks right in the Lobby that opens at 5:30 a.m. and the staff, everyone of them, were nice and helpful!! Would definitely recommend and would stay there again and again!! August 6, 2013
Rated 3 out of 5 by jim2014 Good and bad Beautiful hotel. Concierge great. Club room- service poor.Big prices for drinks, slow service, ran out of food. Parking was overwhelmed and inconvenient. Bar was also overwhelmed. Great location. Front desk great. I was really disappointed in the club room for platinums! They need to improve staffing levels when crowded and not price their drinks at the club level higher than the Ritz bar. December 26, 2013
Rated 2 out of 5 by TTRiii Beware of the elevators During our stay the elevators to the upper floors did not work. The staff tried very hard to make up for the inconvenience but it did not really ease the fact that we couldn't get to our rooms at times. June 21, 2014
Rated 3 out of 5 by SGOLD The rooms were dusty The rooms were dusty and dirty, the food was not good, especially lunch for the conference I was attending. June 30, 2014
Rated 3 out of 5 by Timcat Gr8 location Everything is wonderful except for the petite pool. July 4, 2014
Rated 4 out of 5 by Dawn5 Great location This hotel was within walking distance to the French Quarter and Riverwalk. It was true Sheraton standards. Friendly staff, comfy beds, very clean, and safe. June 24, 2014
Rated 4 out of 5 by CADentist Great location, great value We enjoyed our quick trip to NOLA. I've stayed at other hotels on Canal Street, and the Sheraton is a pretty nice place to stay, way better than average. I like the Starbucks coffee in the room (they have a shop downstairs), even though our coffeemaker didn't work well. We didn't receive any new coffee packets on the second day, nor did we receive any shampoo. However, we did on the third day. The beds were comfortable and the sheets were nice. We didn't spend a lot of time in our room. Our neighbors made too much noise for about half an hour around 3:30am, but that's not the hotel's fault (we could have gotten mad at them but didn't want to ruin their fun). I guess you have to expect that in NOLA. Overall, our hotel stay was nice. The bellmen are extremely efficient! Oh, pretty nice view (between other hotels) our room looking onto the Mississippi River, and close to the French Quarter, cable cars, etc. November 4, 2013
Rated 3 out of 5 by Mateo55 Company meeting Hotel was nice but the construction noise was very distracting during our meeting June 30, 2014
Rated 4 out of 5 by LCK72 mrs Excellent people on the phone(specifically, Shannel) and the morning Front desk on June 25. June 29, 2014
Rated 5 out of 5 by Baldwin23 The recent upgrades are wonderful I have not stayed at a Sheraton in a very long time. My group and meetings were taking place at this Sheraton so that is why I was there. I read online about the recent upgrades and believe they were well done. On top of that, during a rare ice storm, the staff(GM) took great care of communicating with me on what was happening and how it would effect my travel and comfort. January 30, 2014


text:
Max Holland: Starts website
The inspiration for the website comes from an essay penned by George Orwell in 1946, just after the end of World War II. In"Politics and the English Language," Orwell observed that"Political language--and with variations this is true of all political parties, from Conservatives to Anarchists--is designed to make lies sound truthful and murder respectable, and to give an appearance of solidity to pure wind."
That sensibility captures the intent of Washington DeCoded, a non-partisan newsletter about the nation's capital that began publishing on-line on 11 March 2007. Washington DeCoded will not toe any partisan or ideological line. Its only purpose will be to try and explain issues of moment to its readers. Sometimes these issues will be the ones commanding headlines, but more often not.
Max Holland is also a contributing editor at The Nation magazine and the Wilson Quarterly, and sits on the editorial advisory board of the International Journal of Intelligence and Counter- Intelligence. His articles have also appeared in The Atlantic Monthly, American Heritage, The Washington Post, The New York Times, Los Angeles Times, The Boston Globe, The Baltimore Sun, Studies in Intelligence, the Journal of Cold War Studies, Reviews in American History, and on-line at History News Network.
In 2001, Holland won the J. Anthony Lukas Work-in-Progress Award, bestowed jointly by Harvard University's Nieman Center and the Columbia University School of Journalism, for a forthcoming narrative history of the Warren Commission, to be published by Alfred A. Knopf. That same year he won a Studies in Intelligence Award from the Central Intelligence Agency, the first writer working outside the U.S. government to be so recognized.
Holland's books include: The Kennedy Assassination Tapes: The White House Conversations of Lyndon B. Johnson regarding the Assassination, the Warren Commission, and the Aftermath (Knopf, 2004); The CEO Goes to Washington: Negotiating the Halls of Power (Whittle Direct Books, 1994); and When the Machine Stopped: A Cautionary Tale from Industrial America (Harvard Business School Press, 1989).
Although articles appearing in Washington DeCoded are copyrighted, they may be freely distributed and/or printed for non-commercial use unless the copyright is held by another party.
Beginning March 2007, Washington DeCoded will feature monthly articles and book reviews about contemporary history, the political economy, presidential commissions, intelligence and counter- intelligence, 9/11, assassinations, the Cold War, archives and archival practices, secrecy, and U.S. government classification policy.
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- Raleigh Trevelyan, Chronicler of a Notable Family, Dies at 91
- Former spokesman of B.C. anti-immigration group wants UBC history prof fired
- Harvard's Steven Shapin Wins History of Science Award
- Middle East Studies Association Fights a Rising Tide of Critics
- Juan Cole says the postwar Middle East governments were modeled on the Soviet Union, though not communist (interview)

subset: commoncrawl


text:
The Dead Files: A Banshee's Cry Pictures
Steve and Amy investigate reports of violent paranormal activity at Smalley's Inn & Restaurant in upstate New York.Filed under:
Steve talks to a contractor in the basement of Smalley's Inn about why he walked off the job.
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text:
Patrick Leahy
Kirsten Gillibrand, interrupted
Sotomayor's first moment in national spotlight was short and sweet. Kirsten Gillibrand's was neither.
Leahy cuts Gillibrand off
Leahy interrupts Gillibrand's statement, where she heaped lavish praise on Sotomayor.
Sotomayor goes low-key on Day One
Sessions slams "shocking and offensive" belief.
Sunday talk show tip sheet
As the Judiciary Committee gears up for Sotomayor's hearing, a number of senators head to the Sunday talk shows.
Leahy pushes Sotomayor's credentials
He's bringing law enforcement officials to support her.
Tempers flare over Sotomayor
Partisan tension and distrust boil over in the Senate during the past week.
Get thee to the floor -- and fast
The item on Hoyer's agenda with the least chance of getting enacted in the 111th is tardiness reform.
Sotomayor scheduling riles GOP
Leahy pushes past Republicans.
No agreement on Sotomayor timeline
GOP-ers want to review federal cases, mount opposition.
Sotomayor pays a visit to 'SCOTUS Inc'
The business of filling Supreme Court vacancies has become a cottage industry on Capitol Hill.
Leahy wants Sotomayor hearings before August
Judiciary chairman's push threatens GOP go slow strategy.
GOP backs off 'racist' accusations
Top GOP senators try to distance selves from Limbaugh, Gingrich claims that Sotomayor is racist.
Sunday talk show tip sheet
The debate over Supreme Court nominee Sonia Sotomayor dominates talk this Sunday.
Who could lose in confirmation vote?
Which senators will wind up as damaged goods during the confirmation process?
Schumer: Oppose her at your peril
Schumer says the confirmation process will be a "test for the Republican Party."
Leahy: Pelosi not the issue
Leahy accuses Republicans of setting up a "straw woman" to change torture debate focus.
Souter told Leahy of plans in March
The Vermont Democrat says he's known of Souter's decision to retire "for a couple of months."
Obama urged to look beyond courts
Two top senators say Obama should look outside courts when selecting a replacement for Souter.


text:
A method of preparing an electrophoretic support comprising washing of at least a portion of the surface of a silicon-containing support member supporting an electrophoretic matrix with a weak alkali solution and supporting of said matrix by said support member; an electrophoretic gel comprising a polyacrylamide polymer obtained by polymerizing acrylamide or a derivative thereof in the presence of two or more polar organic solvents; a method of electrophoresis employing a gel prepared by said preparation method; and a method of electrophoresis employing said electrophoretic gel.
1. A method for preparing an electrophoretic separation article comprising:
i) providing a support comprising a silicon-containing material and washing at least a portion of a surface of said support for contacting an electrophoretic matrix with a weakly alkaline solution; and
ii) filling or coating said support with an electrophoretic matrix;
thereby obtaining an electrophoretic separation article;
in which the electrophoretic matrix is a composition comprising acrylamide or a derivative thereof or a polymer thereof, and at least two organic solvents.
2. An electrophoretic gel comprising a polyacrylamide polymer or a polymer comprising an acrylamide derivative, and two or more organic solvents, in which one of the organic solvents is formamide.
3. The electrophoretic gel of
4. The electrophoretic gel of
5. The electrophoretic gel of
6. The electrophoretic gel of
7. A process for preparing an electrophoretic gel comprising polymerizing a composition comprising acrylamide or a derivative thereof in the presence of at least two organic solvents.
8. The process of
9. The process of
10. The process of
11. The process of
12. The process of
13. The process of
14. The process of
15. A process for electrophoretic separation of a sample comprising applying said sample to an electrophoretic separation article comprising an electrophoretic gel comprising a polyacrylamide polymer or a polymer comprising an acrylamide derivative, and two or more organic solvents, and separating the sample by applying an electric field electrophoretic separation article.
16. The process of
17. The process of
18. The method of
19. The method of
20. The method of
This application is a Continuation of PCT International Application No. PCT/JP00/06247 filed on Sep. 13, 2000, which was not published in English and which designated the United States and on which priority is claimed under 35 U.S.C. � 120, the entire contents of which are hereby incorporated by reference.
The present invention relates to a method of preparing an electrophoretic support, an electrophoretic matrix, and a method of electrophoresis. More particularly, the present invention relates to a method of preparing an electrophoretic support in which an electrophoretic matrix such as a gel or an entangled polymer is formed on a silicon-containing support member, a method characterized by how the support member is cleaned, and a method of electrophoresis employing the support prepared by this method. The present invention further relates to an electrophoretic gel employed to separate nucleic acids or PNA fragments and to a method of separating nucleic acids or PNA fragments employing this gel or this support.
The method of preparing an electrophoretic support of the present invention inhibits the generation of bubbles during the formation of electrophoretic matrices and thus improves sample (for example, nucleic acid) separation performance. Further the electrophoretic method employing the electrophoretic support or gel of the present invention increases separation performance in electrophoresis. Thus, the separation of long strands of nucleic acid and PNA fragments required by sequencing methods in particular is readily performed, and the present invention is particularly useful when applied to methods of long-strand nucleotide sequencing.
Analysis of the nucleotide sequence of the human genome, said to comprise as many as three billion base pairs, is progressing. In particular, the analysis of polymorphism in the human genome relates to the unique traits of each individual and is the subject of considerable interest in the fields of medicine, pharmacology, and biology. Conventionally, the development of a sequencer capable of automatically processing multiple samples simultaneously, rapidly, and with high sensitivity has been conducted to determine the huge nucleotide sequence of the human genome. In particular, the arrival of multicapillary DNA sequencers simultaneously employing multiple capillary columns packed with gel instead of the tabular slab gel that was previously employed has greatly contributed to increasing the speed of nucleotide sequencing. Currently, 96 capillary column (for example, the Sequencer 3700 from ABI and the MegaBACE 1000 from Molecular Dynamics), and recently, the fourfold 384 capillary column (the development of the 384 multicapillary sequencing system: Proceedings of the 21st Meeting of the Japan Molecular Biology Society, 1P-570 (Yokohama, December 1998) multicapillary DNA sequencers have been developed.
However, when the number of capillary columns is increased, there are limitations due to the performance and structure of the detection devices used to read the sequences. Further, merely increasing the number of capillary columns does not diminish the labor required to supply the capillary columns themselves and to load specimens into capillary columns.
Further, in currently employed sequencers, about 500 bases are read by a single capillary column. Increasing the number of bases read by a single capillary column to increase separation performance has the advantage of reducing the effort required to supply the capillary columns themselves and to load specimens into the capillary columns. However, almost no studies have been conducted into reading more bases at once in individual capillary columns; this remains a problem to be solved in the future.
For example, in many electrophoretic devices including DNA sequencing devices, a support member is employed to support the electrophoretic matrix. The support member has various shapes depending on the electrophoretic objective. For example, in slab electrophoresis, two pieces of flat tabular silica glass are employed. In capillary electrophoresis, a column-shaped (hollow tubular) silicate capillary column is employed as the support member. Further, in micro electrophoresis, a support member with minute separation passages formed on a silicon wafer is employed. All of these support members are generally made of silicon-containing materials, such as silica.
To obtain good separation performance in electrophoretic matrices prepared on such silicon-containing support members (referred to hereinafter as simply �support members�), it is desirable for bubbles not to be present in the matrix.
For example, when electrophoretic gel is sandwiched between two support members (in slab electrophoresis), when an entangled polymer solution is injected into a cavity in a support member (capillary electrophoresis), or when a support member is employed without being cleaned, particularly when the width of the support member or injection inlet are narrow (when the width is less than about 1 mm in diameter), air bubbles tend to develop near the surface and in the minute voids of the support member. Gel and entangled polymer matrices containing such bubbles do not provide adequate separation performance and cannot perform the original functions of electrophoretic matrices.
The method of admixing propanol or polyethylene glycol in advance to polymerization has been proposed as a method of preventing the development of such bubbles (Anal. Chem., 1992, 64, pp. 2665-2671; J. Chromatogr., 1991, 550, pp. 823-830). This method yields a matrix that does not contain bubbles. However, the support obtained by this method has the drawback of having lower separation performance in electrophoresis than conventional supports. Thus, this method is unsuitable for electrophoresis requiring high separation performance.
Further, assuming that impurities present on the surface of the support member are the cause of the bubbles, attempts at inhibiting the occurrence of bubbles have been made by the method of cleaning away impurities on the inner walls of capillary columns using a strong base solution such as NaOH, organic solvents such as ethanol, acid solutions such as HCl, and solutions of these compounds in combination by using pure water (Electrophoresis 1996, 17, pp. 144-151). However, although this method prevents the occurrence of bubbles at the surface of the support member, electrophoretic separation performance is unsatisfactory in the same manner as in the above-described method.
Accordingly, there has been for some time a need to provide a method of preparing electrophoretic supports capable of effectively inhibiting the occurrence of bubbles in matrices carried by or packed in support members that deliver good separation performance.
Further, in addition to methods of preparing electrophoretic supports, there has also been room for improvement in the electrophoretic matrix itself to improve electrophoretic separation performance, However, there has not been adequate investigation of electrophoretic matrix materials and the like.
Accordingly, the object of the present invention is to provide a means of solving the above-stated problems in electrophoresis, improving electrophoretic separation performance, improving sample separation, and particularly, in nucleic acid and PNA fragments, reading even longer-strand nucleotide sequences.
More specifically, the object of the present invention is to provide a method of preparing an electrophoretic support capable of fin inhibiting the generation of bubbles in the matrix when preparing an electrophoretic support employing a silicon-containing support member, and to provide a method of electrophoresis employing the electrophoretic support prepared by this method.
A further object of the present invention is to provide an electrophoretic gel tending not to undergo compression in the nucleotide sequencing of long-strands exceeding 500 bases, and to provide a method of electrophoresis capable of reading long-strand nucleotide sequences using this gel.
The present inventors conducted extensive research into methods of preparing supports capable of improving separation performance without generating bubbles in the course of supporting (carrying or filling) a matrix such as a gel or a polymer with a silicon-containing support member such as a silica capillary column.
As set forth above, the generation of bubbles in the support can be inhibited by cleaning the support member with a strong alkali solution, but despite the inhibition of the formation of bubbles, separation performance does not improve. The present inventors presumed that it was caused by the support member surface being etched by the action of the strong alkali reagent, creating irregularities in the surface of the support member. As a result, dispersion in directions other than the direction of separation of electrophoresis occurs, or the number of free silanol moieties increase at the surface of the support member, preventing improvement in separation performance in electrophoresis.
Accordingly, the present inventors, based on these presumptions, examined various methods in which the etching of the surface of the support member tended not to be etched and the generation of bubbles tended not to occur in the matrix. As a result, they discovered that bubbles are not generated in a matrix that is supported (carried or filled) by a support member that has been cleaned with a weak alkali solution, and that separation performance during electrophoresis improves relative to conventional methods employing strong alkali solutions; the present invention (method of preparing an electrophoretic support) was devised on that basis.
That is, the first mode of implementing the present invention relates to a method of preparing an electrophoretic support wherein at least one portion of the surface of a member (hereinafter �support member�) for supporting an electrophoretic matrix and coming into contact with said matrix is washed, and said matrix is then supported by said support member, characterized in that said support member comprises a silicon-containing material and at least a part of said washing is conducted with a weak alkali solution.
The second mode of implementing the present invention relates to a method of electrophoresis employing an electrophoretic matrix prepared by the above-described method of preparation.
Further, the present inventors investigated the separation of longer strands of nucleic acid or PNA fragments than in the past at one time in one carrier column to a degree permitting nuclectide sequencing, and as a result, improvement in the composition of the electrophoretic gel in a manner permitting nucleotide sequencing was made, as well as the electrophoretic conditions employed when using this gel, and, in particular, electrophoretic solutions were investigated.
Polyacrylamide gel is generally employed in electrophoresis for nucleotide sequencing (reference: PACE (polyacrylamide gel electrophoresis), Hirokawa Shoten, 1990, ed. by Toshio TAKAGI). The present inventors discovered that a gel obtained by preparing a polyacrylamide gel in the presence of two or more polar organic solvents, such as methanol and formamide, solved the above-stated problems; the present invention (electrophoretic gel) was devised on this basis.
That is, the third mode of the present invention relates to an electrophoretic gel comprising a polyacrylamide polymer obtained by polymerizing an acrylamide, or a derivative thereof, in the presence of two or more polar organic solvents.
The fourth mode of the present invention relates to a method of separating nucleic acids or PNA fragments by electrophoresis in the presence or a polar organic solvent.
A highly flexible gel obtained by polymerization in the presence of a water-soluble polymer such as dextran can be employed as the gel of the third mode of the present invention (Katsunori AIZAWA, Tanpakushitu-kakusan-koso 43 (1998) 2191-2198).
Most Preferred Modes of Practicing the Present Invention
Method of Preparing an Electrophoretic Support (Mode 1)
In the method of preparing an electrophoretic support of the present invention, a support member comprising a silicon-containing material is employed to support the matrix. The support member employed here is not specifically limited as to material so long as it contains at least silicon and can be employed in electrophoresis. Examples of the materials of such silicon-containing support members are natural and synthetic silica, borosilicate glass, alkali lead glass, soda lime silica glass and other glass products, quartzite bricks, zeolite, cordierite, silicon carbonate, silicon, enamel, and other compounds chiefly comprised of silicon.
The shape of the support member employed in the present invention may be suitably determined based on what is to be separated. Examples of shapes are tabular, columnar (hollow cylinder), granular, fiber-like, and any combination thereof. Further, any wafer of any shape functioning as a chip in microchip electrophoresis and nanochip electrophoresis is covered by the support member of the present invention.
The diameter and length of the support member may be suitably determined based on the object of separation. For example, in the case of a capillary column, an outer diameter of 100-400 μm, an inner diameter of 2-100 μm and a length of about 10-100 cm are appropriate. When conducting measurement with fluorescent light with a sequencer in capillary columns, it is desirable to employ a material or a support member that has been processed (coated with a nonfluorescent substance) so as not to interfere with fluorescent measurement.
The surface of the support member of the present invention coming into contact with the electrophoretic gel is washed with a weak alkali solution. The weak alkali solution employed in washing is a liquid solution with a solute in the form of a weak alkali substance (including organic substances and inorganic substances), preferably one dissolved in water.
Examples of modes of the weak alkali solutions are organic solutions, inorganic solutions, and mixtures comprising any combination of organic solution and inorganic solution.
Examples of organic solutions are solutions comprising an organic compound, or a combination of multiple organic compounds, selected from the group consisting of aminobutanoic acid, 2-amino-2-ethyl-1,3-propanediol, 2-amino-2-methyl-1,3-propanediol, 2-amino-2-methyl-1-propanol, aminopyridine, butylamine, chloridine, diethanolamine, diethylamine, diethylbarbituric acid, dimethylamine, dimethylimidazole, ephedrine, ethanolamine, ethylmorpholine, glycylglycine, hydroxypyroline, piperidine, propylamine, methylamine, methylimidazole, triethylamine, triethanolamine, trimethylamine, tris(hydroxymethyl)aminomethanol dissolved in a solvent. For example, when dissolved in a solvent at room temperature, the pK value is 7-12 and the pH value is 7-12, preferably 8-11.
Further examples of organic solutions in addition to the above are solutions of organic compounds such as amines, amides, imides, ammonia compounds, and amino acids, and solutions in ammonia water and �Good� buffer solutions, where a slightly alkaline pH is exhibited when the solutions are prepared, as well as solutions adjusted to slight alkalinity with a reagent.
Examples of inorganic solutions are substances selected from among the group consisting of phosphoric acid, pyrophosphoric acid, boric acid, and carbonic acid, exhibiting a pK of 7-12 at room temperature, and when the inorganic compounds are in solution, a weakly alkaline pH (a pH of 7-12, preferably 8-11); solutions prepared to be slightly alkaline with reagents are also included. The inorganic solution of preference is an aqueous solution of carbonate.
The weakly alkaline solution employed in washing in the present invention may also comprise any concentration of mixed-in organic solvent.
In the washing of the present invention, at lease one part of the washing is conducted with an alkaline solution. That is, in the washing of the present invention, it is possible to use other solutions in combination with the weakly alkaline solution. Examples of other solutions that may be employed in combination are: pure water (milli-Q processed pure water) and other neutral solutions; methanol, ethanol, and other organic solvents; and solvents obtained by combining a weakly alkaline solution and a weakly acid solution. The order of washing when employing a solution in addition to the weakly alkaline solution is not specifically limited so long as the washing with a weakly alkaline solution is not omitted; washing with a weakly alkaline solution may be conducted at any step such as first or last. The washing with a solution other than a weakly alkaline solution may be omitted for any reason.
Any time falling within several seconds to several months may be selected as the washing time. However, in consideration of efficiency of operation and the like, a washing time of about several minutes is desirable. The temperature during washing is not specifically limited so long as the solution does not solidify, volatize, or boil away. Washing at room temperature is desirable from the perspective of facilitating the operation.
In the present invention, at least a portion of the surface of the support member coming into contact with the electrophoretic matrix is washed. Since the washing of the support member is conducted with the object of inhibiting the formation of bubbles during preparation of the electrophoretic matrix, the entire surface of the support member coming into contact with the electrophoretic matrix is desirably washed.
Matrices generally employed in electrophoresis can be employed without alteration as the electrophoretic matrix coming into contact with the support member. The matrix may comprise a gel or an entangled polymer, but is not specifically limited thereto. Examples of matrices are polyacrylamide, agarose gel, and entangled polymers having a molecular sieving function such as straight-chain polyacrylamides, water-soluble cellulose derivatives, and dextran. The electrophoretic gel of the present invention that is described as such a matrix further below may also be employed. The use of this gel is suited to the separation of nucleic acids and PNA fragments.
In addition, the methods and conditions that are generally employed for matrices can be employed as the methods and conditions for making and filling matrices in the preparation of electrophoretic matrices coming into contact with the support member. Further, when the matrix is the electrophoretic gel of the present invention, described further below, it can be prepared according to the conditions described further below.
Method of Electrophoresis Employing a Matrix Prepared by the Preparation
Method of the Present Invention (Mode 2)
The present invention covers the method of electrophoresis employing a matrix prepared by the preparation method of the present invention. Based on this method of electrophoresis, various substances can be separated based on the matrix employed in electrophoresis. Examples of such substances are water-soluble substances comprising one, two, or more cationic and/or anionic charges, neutral substances not having a charge, and any mixture thereof. Nucleic acids (DNA and RNA) and PNA fragments are included therein. Additionally, organic substances, inorganic substances (including metal ions), and substances comprising combinations thereof are also included in the above-mentioned substances. A suitable form of the substance to be separated such as a solid, slurry, powder, or liquid may be suitably selected based on the matrix and electrophoresis conditions.
The conditions of electrophoresis such as the buffer solution, voltage, and time employed in the method of electrophoresis of the present invention are suitably selected based on the characteristics of the matrix and the substance to be separated. The conditions of electrophoresis employed when the matrix is the electrophoretic gel of the present invention described further below may also be employed.
Electrophoretic Gel (Mode 3)
The electrophoretic gel of the present invention comprises a polyacrylamide polymer obtained by polymerizing acrylamide or a derivative thereof in the presence of two or more polar organic solvents. Examples of the two or more polar organic solvents employed in the present invention are: methanol, ethanol, 1-propanol, 2-propanol, isobutanol, t-butanol, and other alcohols, pyridine, tetrahydrofuran, dioxane, acetone, acetonitrile, dimethylformamide, dimethylacetamide, dimethylsulfoxide, N-methylpyrrolidone, and hexamethylphosphamide.
The electrophoretic gel of the present invention is obtained by polymerizing an acrylamide derivative in the presence of the polar organic solvents. When an acrylamide derivative is polymerized and two or more polar organic solvents are similarly provided in the gaps of the polymer obtained, the effect of the object of the present invention is not achieved. Nor is the effect of the object of the present invention achieved when only one polar organic solvent is employed in polymerization.
Although the combination of the two or more polar organic solvents employed is not specifically limited, a preferred example is a combination of formamide and an alcohol, preferably the combination of formamide and methanol.
Although the acrylamide derivative employed in polymerization is not specifically limited, examples are N,N′-dimethylacrylamide and N-(hydroxymethyl)acrylamide. The acrylamide derivative may be employed singly or in combinations of one or more. Polymerization initiators that are commonly employed in the polymerization of acrylamide derivatives may be employed without alteration. The concentration (W/V %) of the gel may be suitably determined by considering the molecular weight and the like of the nucleic acid to a be separated. However, it is normally set within a range of 3-10 W/V %, preferably about 5 W/V %.
The method of manufacturing the electrophoretic gel of the present invention is described in greater detail below.
The preparation of a gel at a scale of 100 ml using methanol and formamide as the polar organic solvent will be described. A flask or beaker equipped with a stirring bar is prepared. First, 36 g of urea to make a final concentration of 6 M, 25 ml of pure water (preferably milli-Q processed pure water), 15 ml of �10 tris-boric acid-EDTA buffer solution (referred to hereinafter as �TBE�, in this case �10 TBE), and 10 ml of acrylamide derivative in the form of Long Ranger (U.S. firm FMC) are sequentially added and stirred at room temperature until the urea dissolves. The urea is added as a denaturing agent for nucleic acids or the like; it may be omitted. Urea can be added within a concentration range of 0-8 M. When the concentration of urea exceeds 6 M, the presence of polar organic solvents tends to make the urea precipitate out at low temperatures (0-10� C.). Next, while continuously stirring, the polar organic solvents in the form of 10 ml of methanol and 10 ml of formamide are admixed to adjust the final quantity to 100 ml and stirring is continued. In the present example, the final concentrations of methanol and formamide are 10 percent each. However, the concentration of the polar organic solvents is not limited thereto and may be suitably determined in consideration of the characteristics of the nucleic acid to be separated or the like. For example, a range of 5-15 percent may be set. Further, the concentration of the two or more polar organic solvents do not have to be identical. They may be suitably varied.
As needed, a water-soluble polymer in the form of dextran or some other cellulose derivative may be added to the electrophoretic gel of the present invention. The addition of water-soluble polymers increases the flexibility of the gel (Katsunori ATZAWA, Tanpakushitu-kakusan-koso 43 (1998) 2191-2198). The water-soluble polymers are added in a quantity of 1-30 (W/V) %, preferably about 2-5 (W/V) %. The water-soluble polymers are added to the reaction solution obtained above and suitably stirred to uniformly disperse and dissolve them into the reaction solution.
Minute debris in the reaction solution obtained above is desirably removed. This is because failure sometimes occurs due to the presence of minute debris in electrophoresis in capillary columns. For example, to remove the minute debris in the reaction solution, suction filtration employing a filter (0.22 microns) can be used. However, this is not a limitation.
Once impurities have been removed, the reaction solution is cooled with ice with stirring to inhibit polymerization of the gel. A low temperature chamber may be employed in place of cooling with ice. Once thorough cooling has been confirmed, a suitable amount of ammonium persulfate solution is added as a polymerization initiator. Degassing is conducted as necessary. About 0.5 ml of a 10 percent solution of ammonium persulfate solution can be added. However, this is not a limitation. Degassing of the reaction solution can be conducted, for example, for about 30 min at reduced pressure while stirring the reaction solution. Once the ammonium persulfate solution has been added and degassing has been conducted as necessary, 0.05 ml of a polymerization accelerator in the form of N,N,N′,N′-tetramethylethylenediamine (�TEMED� hereinafter) is added to the reaction solution. Once the TEMED has been added, degassing is desirably conducted for another 5-10 min with stirring. At the point in time where the polymerization accelerator is added, polymerization begins rapidly and gelling commences. Accordingly, conducting the above-described steps with ice cooling or in a low-temperature chamber as set forth above is desirable from the viewpoint of inhibiting gelling prior to packing capillary columns or the like.
Preparation of the Support Member
The material, diameter, length, and the like of the support member employed in the electrophoretic gel of the present invention, as is the case for the support member employed in the above-described method of preparing an electrophoretic support, are not specifically limited. For example, support members commonly employed in multicapillary DNA sequencers may be employed. Additionally, slab gel type support members may be employed. The interior surface of the support member is preferably prepared according to the method of preparing an electrophoretic support of the present invention prior to use.
Packing of Gel into Support Member
The packing of the support member (capillary member) with the reaction solution in which gelling has been started by the addition of the polymerization initiator can be done with a gel packing device such as a GVT unit (from Shimadzu corporation) at room temperature. However, the type of gel packing device is not limited thereto. When a GVT unit is employed as the gel-packing device, the unit is put into pressure mode and all of the capillary columns are filled with solution (reaction solution that has started gelling). The filling of the capillary columns with the gelling solution is confirmed and one to two minutes later the pressure mode is released. The capillary columns that have been filled with gelling solution are left standing for about 3 hours, for example, at room temperature to complete polymerization, after which they are ready for use.
In the case of a slab gel, the same method as is conventionally employed is used to add polymerization initiator, starting gelling. The reaction solution is then packed into the support member (gel plates) and left standing for a prescribed period at room temperature to complete the manufacturing process.
Method of Electrophoresis Employing the Gel of the Present Invention (Mode 4)
The present invention further covers the method of electrophoresis in which nucleic acid or PNA fragments are separated in the presence of polar organic solvents using the support (gel) prepared by the method of preparation of the present invention or the above-described gel of the present invention. The nucleic acid that is separated may be either RNA or DNA. The gel employed in the method of electrophoresis employing the gel of the present invention may also be the matrix prepared by the method of preparation of the present invention (Mode 1). Electrophoresis employing as gel the matrix prepared in this manner is preferable because it permits the separation of longer strands of base pairs.
For example, TBE or the like to which has been added a polar organic solvent is employed as the buffer solution in electrophoresis. Examples of TBE are 0.5-5�, preferably a final concentration of 1.5�. There are no specific limitations on the type, combination, or quantity of polar organic solvent. However, as in the case of manufacturing gel, the use of two or more polar organic solvents is preferred. More preferably, a mixed solution of an alcohol and formamide, and still more preferably, a mixture of methanol and formamide is employed. The concentration of the polar organic solvent is, for example, 5-15 percent, preferably 10 percent (V/V). When using two or more polar organic solvents, the final concentration of each of the solvents is 5-15 percent, preferably 10 percent (V/V). The injection of the sample can be conducted under the same conditions as are normally employed in electrophoresis. For example, in the case of capillary columns, 5-600 seconds at 0.1-10 kV, preferably 90 seconds at 2 kV, is suitable. Migration is suitably conducted at 20-70� C., preferably 55� C., and 1-20 kV, preferably 4.8 kV. In the case of a slab gel, as well, loading can be performed by the usual methods.
Detection of the Separated Matter
Samples that can be separated by an electrophoresis method employing the electrophoretic gel of the present invention are desirably PNA fragments or nucleic acids such as DNA or RNA that have been fluorescent-labeled with a rhodamine compound so as to permit the ready detection of separated matter. However, there is no limitation to such samples; samples labeled with radioactive isotopes may also be separated. The bands of fluorescent-labeled DNA and RNA can be detected using the laser light source and fluorescent light detector of the capillary sequencer. The light source and detector of the sequencer can be employ for slab gels, as well.
The present invention is further described below through embodiments.
The individual resolution (Rs) in the embodiments was calculated using the signal wavelength as the Gauss pattern from the following equation using separation data.
Here, tA and tB denote the times at which the peak A and B signals were detected and WA and WB denote the signal widths at half the peak height of the A and B signals, respectively.
A gel matrix (100 ml) was prepared by sequentially mixing together while continuously stirring at room temperature the following reagents:
The above are admixed, the volume is adjusted to 100 ml with milli-Q pure water, and thorough stirring is conducted again. Filtering (0.22 micron) is conducted at room temperature under negative pressure cooling with ice (10 min) is conducted with stirring, and the solution is left standing in a low-temperature chamber (30 min).
While stirring the solution with ice cooling or in a low-temperature chamber, the stated quantity of a newly prepared batch of this reagent is added. Degassing is then conducted for 30 min with stirring.
This reagent is added quickly and further degassing is conducted for about 10 min with ice cooling or in a low-temperature chamber with constant stirring.
The gel of the solution thus obtained was packed at room temperature into capillaries (SGE Co., internal diameter 100μ, external diameter 360μ, made of fused silica, total length 48 cm) using a GVT unit (Shimadzu Corporation). After packing, the capillaries were left standing for 3 hours at room temperature, yielding capillary columns filled with the gel of the present invention. These capillary columns were then employed in electrophoresis.
Sample Injection
DNA fragments that have been fluorescence-labeled by cycle sequencing or the like are purified by desalting or ethanol precipitation. Formamide or a mixed solution of formamide and buffer solution is admixed to the sample and high-temperature processing (95� C., 2 min) is conducted. A voltage was applied for from 5 seconds to 5 minutes at a range of 5-100volts per centimeter of length of the capillary to inject the sample into the capillary through the capillary inlet on the cathode side.
The following components were continuously stirred at room temperature to prepare the electrophoretic buffer solution.
An argon laser was directed through a detection window positioned near the anode side of the capillary, the fluorescent agent in the DNA sample was excited, and the fluorescent light thus generated was detected with a photo-multimeter.
Table 1 compares the conditions under which the polar organic solvents were present during gel production and improvements in the ability to read nucleotide sequences of the gels. As is evident from the embodiment employing formamide and methanol, at least two polar organic solvents were required; the effect was not achieved with just one. Further, to the extent investigated, the optimal concentration of the two organic solvents was 10 percent (v/v) each. Under these conditions, it was sometimes possible to read 900 bases or more.
Table 2 shows whether or not a polar organic solvent was present during gel production, whether or not a polar organic solvent was present in the buffer solution used in electrophoresis, and how the ability of the gels to read nucleotide sequences was affected. As is clear from these tables, the best results were achieved when the organic solvents were present both during gel preparation and during electrophoresis.
Support Member Employed
In the method of preparing an electrophoretic support of the present invention, 384 fused silica capillary columns from Moritex Co. were employed as the support member. These capillary columns were 48 cm in length, had a length of 39.5-40 cm from inlet to detection window, measured 100 μm in inner diameter and 300 μm in outer diameter, and had uncoated internal surfaces.
Washing was sequentially conducted at room temperature with 0.1 M NaHCO3 (pH 9.0), pure water (milli-Q processed water), 1 M HCl, and pure water (milli-Q processed water). For comparison, washing was also sequentially conducted by the conventional method with 1 M NaOH (pH 13-14), pure water (milli-Q processed water), 1 M HCl, and pure water (milli-Q processed water). Washing with each solution lasted for 3 minutes.
Preparation of Matrix (Gel) and Injection into Column
Gel matrices (100 ml) were prepared in 385 of the above-described washed fused silica capillary columns
First, acrylamide/bis, the commercial product Long Ranger (U.S. FMC Co.), or Page Plus (Amresco Co.) monomer solution were used to prepare the gel matrices. The composition of the solution prior to polymerization was as follows.
A quantity of TEMED yielding a final concentration of 0.06 percent (V/V) was added to the solution of the above-stated composition prior to polymerization and degassing was conducted for 30 min while cooling with ice. A quantity of ammonium persulfate yielding a final concentration of 0.05 percent (W/V) was then added and the solution was further degassed for 5 min.
At the point when the polymerization reaction began, each of the 384 capillary columns was filled with gel by the same method as in Embodiment 1 and the capillary columns were placed at room temperature.
Separately from the above-described gel preparation, standard samples for sequencing (pGEM-3Zf(+)/21M13 forward primer) and fluorescent-labeling agent (BigDye terminator) were prepared.
Sequencing (electrophoresis) was conducted with the gel-filled capillary columns and a RISA sequencer (Shimadzu Corporation) at an applied voltage of 4.8 kV and a temperature of 50� C. The conditions were identical to those in Embodiment 1
As shown in Table 3, when the gel comprised Page Plus, for example, and washing was conducted with 0.1 M NaHCO3, 750 nucleotide sequences were read at a resolution per base of 0.5. This roughly matches the number of nucleotide sequences that can be read with a RISA sequencer. By contrast, when washing was conducted with 1 M NaOH, only 560 nucleotide sequences could be read at a resolution per base of 0.5. Comparing gels of solutions of identical composition prior to polymerization shows that regardless of the type of gel, washing with 0.1 M NaHCO3 permitted the reading of longer strands of nucleotide sequences than when washing was conducted with 1 M NaOH.
Further, when a single washing solution was employed, gels containing both formamide and methanol permitted the reading of longer strands of nucleotide sequences than gels that did not contain formamide and methanol.
Further, even when washing was conducted with either 0.1 M NaHCO2 or 1 M NaOH, gels containing both formamide and methanol permitted the reading of longer strands of nucleotide sequences than gels that did not contain both.
From the results of the presence of both formamide and methanol in the gels, it was found that the use of 0.1 M NaHCO3 had a synergistic effect on washing.
No bubbles were seen for any type of gel in the capillary column when processed with 0.1 M NaHCO3. When 0.1 M NaHCO3 (pH 9.0) was employed in washing, safety increased for work factors such as handling relative to the use of 1 M NaOH.
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|By Peter Silva
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|February 21, 2013 09:00 AM EST
BYOD has quickly transformed IT, offering a revolutionary way to support the mobile workforce. The first wave of BYOD featured MDM solutions that controlled the entire device. In the next wave, BYOD 2.0, control applies only to those apps necessary for business, enforcing corporate policy while maintaining personal privacy. The #F5 Mobile App Manager is a complete mobile application management platform built for BYOD 2.0.
As more smartphones, tablets, and other types of mobile devices make their way into employees' hands, requests for corporate access from those devices are increasing, which represents a huge challenge for IT departments. Not only has IT lost the ability to fully control and manage these devices, but employees are now demanding that they be able to conduct company business from multiple personal devices. Initially resistant to the idea due to security concerns, IT teams are slowly adopting the concept, but hesitantly, still concerned about the inherent risks of allowing personal devices to access and store sensitive corporate information.
People have become very attached to their mobile devices. They customize them, surf the web, play games, watch movies, shop, and often simply manage life with these always-connected devices. The flipside of the convenience and flexibility of BYOD are the many concerns about the risks introduced to the corporate infrastructure when allowing unmanaged and potentially unsecured personal devices access to sensitive, proprietary information. Organizations need dynamic policy enforcement to govern the way they now lock down data and applications. As with laptops, if an employee logs in to the corporate data center from a compromised mobile device, then that employee becomes as much of a risk as a hacker with direct access to the corporate data center.
Enter BYOD 1.0.
BYOD 1.0 is the industry's first attempt at solving problems related to personally owned devices in the workplace. BYOD 1.0 consists of two primary components—mobile device management (MDM) and device-level, layer 3 VPNs. The primary goal of MDM is to manage and secure the endpoint device itself, including varying amounts of protection for data at rest on the device (which is typically limited to enabling native device encryption via configuration). The primary aim of the layer 3 VPN is to connect the device back into the corporate network, providing data-in-transit security for corporate traffic.
Both of these BYOD 1.0 components have a drawback—they are umbrellas that protect and manage the entire device, rather than zeroing in on just the enterprise data and applications on that device. Since these are usually dual-purpose (work/personal) devices, this device-wide approach causes issues for both workers and for IT. Employees don't like that BYOD 1.0 imposes enterprise controls over their personal devices, applications, and information. One of the most commonly cited examples is that of the employee who leaves a company and has his device wiped by the organization, losing photos of his family along with the enterprise data and applications. People are also concerned with the privacy of their personal data under a BYOD 1.0 scheme.
From an IT perspective, organizations agree—they don't want to have to concern themselves with personal data or applications. As soon as they manage the entire device or simply connect that device to the corporate network via VPN, that personal traffic also becomes an IT problem. While BYOD 1.0 helps to enable the use of personally owned devices in the enterprise, the device-level approach certainly has its challenges. BYOD 2.0 seeks to solve these shortcomings. The shift from BYOD 1.0 to BYOD 2.0 builds on many of the concepts developed during BYOD 1.0, adding a new set of frameworks that enable IT organizations to wrap enterprise applications in a security layer.
Throughout BYOD 1.0, F5 has provided connectivity for mobile devices into enterprise networks with VPN functionality, most commonly through iOS and Android versions of the F5 BIG-IP Edge Client. This layer provides management capabilities as well as functionality such as authentication and authorization, data-at-rest security, and data-in-transit security, among others.
BYOD 2.0 builds on the BYOD 1.0 foundation but makes a substantial shift from a device-level focus to an application-level focus. BYOD 2.0 seeks to ensure that the enterprise footprint on a personally owned device is limited to the enterprise data and applications and nothing more. This means that mobile device management is supplanted by mobile application management (MAM), and device-level VPNs are replaced by application-specific VPNs. These application-specific VPNs include technology such as BIG-IP APM AppTunnels, a single secure, encrypted connection to a specific service such as Microsoft Exchange.
With this approach, workers are happier than with BYOD 1.0 because the enterprise manages and sees only the enterprise subset of the overall data and applications on the device, leaving the management of the device itself, and of personal data and applications, to the device's owner. IT staff prefer the BYOD 2.0 approach for the same reasons—it allows them to concern themselves only with the enterprise data
and applications they need to secure, manage, and control.
BYOD 2.0 and the aforementioned application wrapping frameworks are changing the dynamic in the mobile space. By combining mobile management functionality and access functionality into a single offering, these wrappers give enterprises a mobile IT solution that extends from data and applications on the endpoint into the cloud and data center.
Introducing F5 Mobile App Manager
F5 Mobile App Manager (MAM) is a mobile application management and access solution that securely extends the enterprise to personal mobile devices. It manages applications and secures data while satisfying the needs of employees and enterprise IT departments. For IT, it limits the burden associated with securing and controlling personal data and mobile use. For employees, it safely separates personal data and use from corporate oversight. F5 MAM is a complete mobile application management platform offering security, management, and compliance for BYOD deployments. It is a true enterprise device, data, and information management solution that fits the needs of the mobile enterprise better than MDM solutions. F5 MAM includes a suite of business productivity applications and capabilities to separate and secure enterprise mobile applications while providing end-to-end security.
F5 MAM Workspace
Organizations and employees both want the ability to segregate professional and personal information. F5 MAM Workspace is an innovative solution allowing enterprises to truly create a virtual enterprise workspace on a wide variety of mobile devices. With MAM Workspace, individuals can have separate sectors and associated policies for their personal and enterprise uses of a device. This enables IT to control how employees access key corporate information while ensuring that employees maintain the freedom to take full advantage of their mobile devices. The secure MAM Workspace can be protected by a password or PIN that is independent of the device password. IT can also reset a user's MAM Workspace password, lock down a user's MAM Workspace, or wipe the Workspace in the event of a policy violation.
F5 MAM App Wrapper
Organizations can also add their own applications to the secure workspace. Organizations have the ability to add any application to the secure, IT-controlled environment. In addition, there is zero need to recompile to create a secure application. F5 MAM App Wrapper scans the existing code in third-party apps, identifies any security vulnerabilities, and injects new proprietary code. This wraps and secures the app for manageability and deployment.
F5 MAM Connect
Email is one of the most critical communication tools for organizations and employees alike. No email, no work. F5 MAM Connect is a secure, wrapped personal information manager (PIM) client that integrates with Microsoft Exchange and delivers enterprise email, calendar, contacts, tasks, and notes to the employee. MAM Connect offers EAS synchronization, global address list integration, secure storage, and networking
and is fully managed via the MAM management console.
F5 MAM Browser
The F5 MAM Browser is a secure and managed browser delivered within MAM. It provides employees with a full-featured browser, separate from their personal browsers, with the control IT needs for secure browser access. It facilitates integrated blocked and safe lists without reliance on proxies, provides controls for enterprise proxy configuration, and allows administrators to push configuration via the web-based MAM portal.
Whether organizations are prepared or not, BYOD is here, and it is transforming enterprise IT. It can potentially provide organizations a significant cost savings and productivity boost, but it is not without risk. F5 provides strategic control points for mobile applications from the endpoint to the data center and to the cloud, enabling unparalleled security, performance, and agility. F5 Mobile App Manager helps organizations make the leap to BYOD or transition from controlling the entire device to simply managing corporate applications and data on the device, solving the work/personal dilemma.
With F5 Mobile App Manager, BYOD 2.0 is now a reality.
- F5′s Feeling Alive with Newly Unveiled Mobile App Manager
- Is BYO Already D?
- Will BYOL Cripple BYOD?
- Freedom vs. Control
- BYOD Uptake Has Only Just Begun
- BYOD Policies – More than an IT Issue Part 1: Liability
- BYOD Policies – More than an IT Issue Part 2: Device Choice
- BYOD Policies – More than an IT Issue Part 3: Economics
- BYOD Policies – More than an IT Issue Part 4: User Experience and Privacy
- BYOD Policies – More than an IT Issue Part 5: Trust Model
Technorati Tags: f5,byod,smartphone,mobile,mobile device,risk,research,silva,security,compliance
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Explosive growth in connected devices. Enormous amounts of data for collection and analysis. Critical use of data for split-second decision making and actionable information. All three are factors in making the Internet of Things a reality. Yet, any one factor would have an IT organization pondering its infrastructure strategy. How should your organization enhance its IT framework to enable an Internet of Things implementation? In his session at Internet of @ThingsExpo, James Kirkland, Chief Architect for the Internet of Things and Intelligent Systems at Red Hat, described how to revolutioniz...
Nov. 21, 2014 09:15 PM EST Reads: 1,401
The security devil is always in the details of the attack: the ones you've endured, the ones you prepare yourself to fend off, and the ones that, you fear, will catch you completely unaware and defenseless. The Internet of Things (IoT) is nothing if not an endless proliferation of details. It's the vision of a world in which continuous Internet connectivity and addressability is embedded into a growing range of human artifacts, into the natural world, and even into our smartphones, appliances, and physical persons. In the IoT vision, every new "thing" - sensor, actuator, data source, data con...
Nov. 21, 2014 08:00 PM EST Reads: 1,404
Technology is enabling a new approach to collecting and using data. This approach, commonly referred to as the "Internet of Things" (IoT), enables businesses to use real-time data from all sorts of things including machines, devices and sensors to make better decisions, improve customer service, and lower the risk in the creation of new revenue opportunities. In his General Session at Internet of @ThingsExpo, Dave Wagstaff, Vice President and Chief Architect at BSQUARE Corporation, discuss the real benefits to focus on, how to understand the requirements of a successful solution, the flow of ...
Nov. 21, 2014 08:00 PM EST Reads: 1,459
"BSQUARE is in the business of selling software solutions for smart connected devices. It's obvious that IoT has moved from being a technology to being a fundamental part of business, and in the last 18 months people have said let's figure out how to do it and let's put some focus on it, " explained Dave Wagstaff, VP & Chief Architect, at BSQUARE Corporation, in this SYS-CON.tv interview at @ThingsExpo, held Nov 4-6, 2014, at the Santa Clara Convention Center in Santa Clara, CA.
Nov. 21, 2014 07:00 PM EST Reads: 1,309
Focused on this fast-growing market's needs, Vitesse Semiconductor Corporation (Nasdaq: VTSS), a leading provider of IC solutions to advance "Ethernet Everywhere" in Carrier, Enterprise and Internet of Things (IoT) networks, introduced its IStaX™ software (VSC6815SDK), a robust protocol stack to simplify deployment and management of Industrial-IoT network applications such as Industrial Ethernet switching, surveillance, video distribution, LCD signage, intelligent sensors, and metering equipment. Leveraging technologies proven in the Carrier and Enterprise markets, IStaX is designed to work ac...
Nov. 20, 2014 09:15 PM EST Reads: 1,389
C-Labs LLC, a leading provider of remote and mobile access for the Internet of Things (IoT), announced the appointment of John Traynor to the position of chief operating officer. Previously a strategic advisor to the firm, Mr. Traynor will now oversee sales, marketing, finance, and operations. Mr. Traynor is based out of the C-Labs office in Redmond, Washington. He reports to Chris Muench, Chief Executive Officer. Mr. Traynor brings valuable business leadership and technology industry expertise to C-Labs. With over 30 years' experience in the high-tech sector, John Traynor has held numerous...
Nov. 20, 2014 06:00 PM EST Reads: 1,349
The 3rd International @ThingsExpo, co-located with the 16th International Cloud Expo - to be held June 9-11, 2015, at the Javits Center in New York City, NY - announces that it is now accepting Keynote Proposals. The Internet of Things (IoT) is the most profound change in personal and enterprise IT since the creation of the Worldwide Web more than 20 years ago. All major researchers estimate there will be tens of billions devices - computers, smartphones, tablets, and sensors - connected to the Internet by 2020. This number will continue to grow at a rapid pace for the next several decades.
Nov. 20, 2014 01:00 PM EST Reads: 1,593
The Internet of Things is not new. Historically, smart businesses have used its basic concept of leveraging data to drive better decision making and have capitalized on those insights to realize additional revenue opportunities. So, what has changed to make the Internet of Things one of the hottest topics in tech? In his session at @ThingsExpo, Chris Gray, Director, Embedded and Internet of Things, discussed the underlying factors that are driving the economics of intelligent systems. Discover how hardware commoditization, the ubiquitous nature of connectivity, and the emergence of Big Data a...
Nov. 20, 2014 12:30 PM EST Reads: 1,804
Almost everyone sees the potential of Internet of Things but how can businesses truly unlock that potential. The key will be in the ability to discover business insight in the midst of an ocean of Big Data generated from billions of embedded devices via Systems of Discover. Businesses will also need to ensure that they can sustain that insight by leveraging the cloud for global reach, scale and elasticity.
Nov. 18, 2014 09:00 PM EST Reads: 2,024
SYS-CON Events announced today that IDenticard will exhibit at SYS-CON's 16th International Cloud Expo®, which will take place on June 9-11, 2015, at the Javits Center in New York City, NY. IDenticard™ is the security division of Brady Corp (NYSE: BRC), a $1.5 billion manufacturer of identification products. We have small-company values with the strength and stability of a major corporation. IDenticard offers local sales, support and service to our customers across the United States and Canada. Our partner network encompasses some 300 of the world's leading systems integrators and security s...
Nov. 18, 2014 08:15 PM EST Reads: 1,583
IoT is still a vague buzzword for many people. In his session at @ThingsExpo, Mike Kavis, Vice President & Principal Cloud Architect at Cloud Technology Partners, discussed the business value of IoT that goes far beyond the general public's perception that IoT is all about wearables and home consumer services. He also discussed how IoT is perceived by investors and how venture capitalist access this space. Other topics discussed were barriers to success, what is new, what is old, and what the future may hold. Mike Kavis is Vice President & Principal Cloud Architect at Cloud Technology Pa...
Nov. 18, 2014 01:30 PM EST Reads: 2,016
Cloud Expo 2014 TV commercials will feature @ThingsExpo, which was launched in June, 2014 at New York City's Javits Center as the largest 'Internet of Things' event in the world. The next @ThingsExpo will take place November 4-6, 2014, at the Santa Clara Convention Center, in Santa Clara, California. Since its launch in 2008, Cloud Expo TV commercials have been aired and CNBC, Fox News Network, and Bloomberg TV. Please enjoy our 2014 commercial.
Nov. 13, 2014 05:00 AM EST Reads: 3,549


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SINGLE FAMILY HOME: 4 bed, 2 full bath, 1 half bath. This meticulously maintained 2 story home on a full partially finished walk-out basement shows likeMore info
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Enjoy the excitement of learning to draw and paint. Basic instruction given in drawing composition, mixing colors, etc. Students to furnish own materials (list available).
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Rabbi Michelle Greenberg will lead a 10:30 a.m. Kabbalat Shabbat service, followed by a hot kosher lunch from Betty the Caterer. The menu will feature: matzah ball soup; stuffed chicken capon with poulet sauce; vegetable blend; Franconia potatoes; challah roll; and birthday cake.
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Exclusive Plus Dream Lodge at Woodlands Park - Westfield, East Sussex
Imagine a haven away from the hustle and bustle of everyday life, a sanctuary far from the stresses of a fast moving world. Set in the heart of the beautiful East Sussex countryside just a few miles from the historic towns of Battle and Rye, Woodlands Park is that place. Less than 60 miles from London, it's a … more


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Adox Color Implosion... What a strange film!
Bought it at the photokina and had to try it the same day. Looks a bit like xpro at bright light and like an overaged film at darker skies... Strange! :-DThis photo was shot on film. Taken by marcel2cv with a Lomo LC-A Refurbished loaded with Adox Color Implosion film in Düsseldorf, Germany. These tags describe this photo: adox, colour_implosion, lomo, lc-a, köln, photokina, and messe. Date: 2012-09-22. Time: Afternoon. This photo can also be found in the album Adox Color Implosion at the photokina.
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text: Snow Girl Snow Globe by Lime Lace £16.95$32.33€22.31$28.37 In stock £2.75$5.24€3.62$4.60 mainland UK delivery - Order now and receive this item on or before Saturday 29 November £2.75$5.24€3.62$4.60 mainland UK deliveryEstimated delivery: 2-5 days £7.70$14.69€10.13$12.89 Express Next Day - Order now and receive this item on or before Wednesday 26 November Express delivery available International delivery available quantity by Lime Lace Location: North Yorkshire, GB read seller's profile see seller's complete range more items from this seller Girls Eskimo Snow Globe £16.95$32.33€22.31$28.37 White Polar Bear Snow Globe £16.95$32.33€22.31$28.37 Kate Bush Album In Book Form Print £24.95$47.60€32.85$41.77 White Glass Fox Decoration £4.95$9.44€6.51$8.28 Personalised Bottle Opener £21$40.06€27.65$35.16 Personalised Mans Copper Cuff £25.95$49.51€34.16$43.44 view all you may also like PRODUCT DETAILS DELIVERY RETURNS ask seller a question Charming snow girl snow globe. This cute snow doll snow globe with falling snow flakes creates a wonderful nostalgic feeling. The cute snow doll is all wrapped up ready for winter in her soft fleecy outfit. Give the globe a little shake to make the snow fall around her and imagine yourself in winter wonderland! made from: Plastic dimensions: H 8cm product code: 284088 This product is dispatched to you by Lime Lace. MAINLAND UK order now and receive this item on or before price Standard Delivery Saturday 29 November £2.75$5.24€3.62$4.60 Two Day Delivery Thursday 27 November £6.70$12.78€8.82$11.21 Express Next Day Wednesday 26 November £7.70$14.69€10.13$12.89 NON-MAINLAND UK estimated delivery price Highlands & Islands of Scotland 2-5 daysfrom dispatch £2.75$5.24€3.62$4.60 Northern Ireland 2-5 daysfrom dispatch £2.75$5.24€3.62$4.60 Other UK Islands 2-5 daysfrom dispatch £4.95$9.44€6.51$8.28 INTERNATIONAL DELIVERY estimated delivery price Western Europe 2-5 daysfrom dispatch £9.95$18.98€13.10$16.65 Rest of Europe 5-10 daysfrom dispatch £9.95$18.98€13.10$16.65 USA & Canada 5-10 daysfrom dispatch £17.95$34.24€23.63$30.05 Far East & Australasia 5-10 daysfrom dispatch £17.95$34.24€23.63$30.05 Rest of the world 5-14 daysfrom dispatch £17.95$34.24€23.63$30.05 You have 28 days, from receipt of cancellable goods, to notify the seller if you wish to cancel your order or exchange an item. Please note: goods that are personalised, bespoke or made-to-order to your specific requirements, perishable products and personal items sold with a hygiene seal (cosmetics, underwear) in instances where the seal is broken are non-refundable, unless faulty. Read more about our returns policy.Should you choose to return or exchange your order you will need to deliver the item(s) to the UK, where this seller is based. Lime Lace's return policy


text: Bajaj Majesty 3 Litres Instant Water HeaterOur Price Rs. 3,300 Add to Compare MRP Rs. 4,000 18Key Features3 Litres Storage Water HeaterThermoplastic Body with ISI MarkMultiple Safety System, 4 Star Rating Store Pickup [?] Buy online, pick up from a Croma Store near you. Get an assured gift coupon on every purchase! Express Delivery [?] Buy online and get your order delivered within 24 hours!Quick View


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Marilyn Manson
Marilyn Manson Denies Involvement in Lana Del Rey Sexual Assault Depiction Footage
Update: According to Marilyn Manson's camp, Manson was not involved with the Del Rey scenes. "Manson did not direct this, shoot it, nor was ...
November 20, 2014 at 10:12 a.m. EST
Die Antwoord Enlist Marilyn Manson, Jack Black, Flea, Cara Delevingne, More for "Ugly Boy" Video
Die Antwoord have shared the video for "Ugly Boy", from this year's Donker Mag. Directed by Ninja and produced by Yo-Landi Vi$$er, it ...
November 4, 2014 at 11:42 a.m. EST
Listen: Girl Talk Mixes Kanye West and Marilyn Manson, Daft Punk and Michael Jackson
When Kanye West debuted "Black Skinhead" on "Saturday Night Live" in May, it sure sounded like the track was sampling Marilyn Manson's 1996 hit ...
July 18, 2013 at 4:18 p.m. EDT
Interrobang (?!)
Interrobang (?!): "Franco-Crunk Fille Feeler-Uppers" b/w "Cuizinier Blew My Mind"
MZ. Julianne Escobedo Shepherd; April 27, 2006
Ceci n'est pas une disque du Southern rap: TTC's Franco-crunk.


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RANCHO SANTA MARGARITA – Members of Girl Scout Troop 447 in Rancho Santa Margarita were recognized by the City Council recently for earning the Bronze Award, the highest honor given a Girl Scout Junior.
The girls received the award after renovating the Cielo Vista Elementary School garden. They removed weeds and old wood chips, planted and cultivated seeds, and added handmade stepping stones and signs.
The city recognized their achievement during Wednesday's council meeting.
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February 7, 2012 | Video
Auckland, New Zealand-based nail tech Leah Light is an international nail correspondant for Perez Hilton's blog. Here, she shows viewers how to create a tuxedo manicure. View more of her videos at http://cocoperez.com/category/leah-light.
February 5, 2012 | Video
For VietSALON Magazine's January/February 2012 cover, Robert Nguyen added a gel overlay to his extreme stilettos and sculpted a dragon and waves with OPI Absolute Crystal Clear acrylic powder.
January 12, 2012 | Video
FingerNailFixer Holly Schippers uses Shellac to create her own nail art decals. You can use any brand of gel polish with this technique.
January 4, 2012 | Video
Nail tech Robin Moses shows us how to create a traditional marbleized nail art design (without using water). See more of her videos at www.robinmoses.com.
To learn how to marble WITH water, here's a tutorial for that:
China Glaze Polish Swirl Live Demo
December 23, 2011 | Video
This difficult-but-worth-it animal print design by Gina Silvestro uses Akzéntz gels and loose glitters.
December 22, 2011 | Video
Gina Silvestro is an Akzéntz educator. Like other great designs for beginners, this design relies on creating simple swirls. They don't have to be perfect, so just pull out a dotting tool and go for it. View more of Silvestro's tutorials at www.gelessentialz.com.
December 7, 2011 | Video
Patrice "Tracie" McNeal of WildSide Nailz Art Studio in Lancaster, Calif., shares her tutorial for a simple way to dress up a French manicure with a little sparkle. Watch more videos by McNeal at www.youtube.com/ladynailz.
December 3, 2011 | Video
Patrice "Tracie" McNeal of WildSide Nailz Art Studio in Lancaster, Calif., shares her tutorial for creating these two-way golden acrylic abalone nails. Watch more videos by McNeal at www.youtube.com/ladynailz.
December 2, 2011 | Video
The folks at Tip Jar, a manufacturer of stylish pre-designed nail tips, show NAILStv users how to easily and correctly apply the tips to clients' nails.
December 1, 2011 | Video
Try making different variations of this design by Lina Nyberg of laLUNE Design, Stockholm, Sweden, by using different colors of papers and cutting out different shapes. If you're familiar with wraps, then this design will be super simple for you.
November 30, 2011 | Video
Maja Rokov of Arty Nails Croatia uses Kinetics products to create this foil flower nail art design.
November 29, 2011 | Video
Maja Rokov of Arty Nails Croatia uses Kinetics products to create this happy floral nail art design.
November 23, 2011 | Video
FingerNailFixer (Holly Schippers) receives lots of requests for foil application demos. Here is a video on how to add transfer foil or metallic leafing to your Shellac.
Holly Schippers (FingerNailFixer) shows you how to take your Shellac for a walk on the wild side with animal print in either leopard, jaguar, cheetah, zebra, or tiger!
November 12, 2011 | Video
Hillary Fry, AKA Solessence, shows us how to do easy glitter nails that look out of this world. This tutorial uses four different glitters, from near-dust particles to hexagonal holo discs to show how you can vamp up a look with little time added to a service. Fry says, "While it may seem redundant to use microfine and ultrafine glitter, it is about the details. On the nail, the microfine adds a sheer base, almost looking like milkyway gases among stars. Experiment with layers to find your favorite looks." Watch more of Fry's videos at www.youtube.com/solessence.
Adore Dolls Parlour Dazzles With Its Nail Art19 photos
Celebrity Face Off Contest: Portrait of an Artist11 photos
Learn More
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People Ain't Isht: Pair Of California Kush Growers Popped For Keeping 15-Year-Old Locked In Metal Box As Sex Slave
Posted on July 26th, 2013 - By Bossip Staff
Categories: Arrests, Kush Chronic-les, News, One-Time, People Ain't Isht, Shady, Skanky, SMH, What the Hell???, What Were You Thinking?, You Can't Be Serious...
Pushing kush is bad enough — but kidnapping and turning out teens is completely disgusting! Continue »


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18326 Apartments for Rent
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"The shooting down of the Malaysia Airlines Boeing 777 in Eastern Ukraine on 17 July has placed the conflict which has engulfed that part of Ukraine into an entirely new context. It has transformed the event from a localised, regional rebellion into a crisis that brings Russia's role into the open, writes Kataryna Wolczuk, of the Centre for Russian and European Studies."
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I think Putin sees himself as a strongman and compared to the wests vaccillating vote-chasing public school puddings, he is absolutely right.
War by proxy is nothing new. In the Korean War, for example, US fighter pilots in 'Mig Alley' knew that they were engaging Russian pilots flying Russian planes in direct combat, notwithstanding the Korean 'flags of convenience'. It hardly mattered at the time, given that the Cold War deadlock was a fait accompli - But it certainly matters now, given Russia's aspirations to international acceptance in trade and relations.
This incident reminds us that NATO is, alas, not such an anachronism.
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