The present invention provides isolated polypeptides comprising an amino acid sequence of a choline binding protein CbpG. This invention provides an isolated polypeptide comprising an amino acid sequence of a choline binding polypeptide CbpG or N-terminal CbpG truncate, including analogs, variants, mutants, derivatives and fragments thereof This invention further provides an isolated immunogenic polypeptide comprising an amino acid sequence of a choline binding protein CbpG. This invention provides an isolated nucleic acid encoding a polypeptide comprising an amino acid sequence of a choline binding protein CbpG. This invention provides pharmaceutical compositions, vaccines, and diagnostic and therapeutic methods of use of the isolated polypeptides and nucleic acids. Assays for compounds which alter or inactivate the polypeptides of the present invention for use in therapy are also provided.

The present invention relates generally to choline binding polypeptides and to nucleic acids encoding such polypeptides. The invention also relates to vaccines which provide protection or elicit protective antibodies to bacterial infection, and to antibodies and antagonists against or inhibitors of such polypeptides for use in diagnosis, therapy and passive immune therapy. In particular, the choline binding polypeptides of the invention are useful as vaccines against Streptococcus, particularly pneumococcus. A choline binding polypeptide of the present invention is also useful as a competitive inhibitor of bacterial adhesion, or to discover small molecule antagonists of adhesion.
Streptococcus pneumoniae is a gram positive bacteria which is a major cause of invasive infections such as sepsis, meningitis, otitis media and lobar pneumonia (Tuomanen et al NEJM 322:1280-1284, 1995). Vaccination has long been an important armament in the arsenal against infectious microorganisms. Prior to the introduction of antibiotics, vaccination was the major hope for protecting populations against viral or bacterial infection. With the advent of antibiotics, vaccination against bacterial infections became less important. However, the emerging problems withantibiotic resistance among such infectious bacteria, including S. pneumoniae strains, have raised an urgent need for a better understanding of the pathogenesis of these pathogens and has reestablished the importance of anti-bacterial vaccines (Appelbaum, P.C. (1992) Clin Infect Dis 15:77-83).
There are over 90 different types of the pneumococcal organism, each with a different chemical structure of the capsular polysaccharide (i.e., each antigenically distinct). The capsular polysaccharide is a principal known virulence factor of the pneumococcus and induces an antibody response in adults. Current vaccines against S. pneumoniae employ mixtures of the capsules of the 23 most common serotypes of this bacterium. After several comprehensive studies there is now overwhelming evidence that this vaccine is approximately 60% efficacious for the general population (Shapiro et al.(1991) NEJM 325:1453-60). Importantly, these vaccines are ineffective in individuals most susceptible to pathological infectionxe2x80x94the young, the old, and the immune compromisedxe2x80x94because of their inability to elicit a T cell immune response. Conjugation of the capsule to a protein permits protection in the younger age group but is inherently limited in the number of capsules that can be conjugated at one time (approximately 5-8 capsules only).
Exported proteins in bacteria participate in many diverse and essential cell functions such as motility, signal transduction, macromolecular transport and assembly, and the acquisition of essential nutrients. For pathogenic bacteria such as S. pneumoniae, many exported proteins are virulence determinants that function as adhesins to colonize and thus infect the host, or as toxins to protect the bacteria against the host""s immune system (for a review, see Hoepelman and Tuomanen (1992) Infect Immun 60:1729-33). One alternative to current vaccines are subunit vaccines in which the antigen, or antigens, include a bacterial surface protein or proteins. These vaccines could overcome the deficiencies of whole bacterial or capsule-based vaccines. Moreover, given the importance of exported or surface proteins to bacterial virulence, these proteins are an important target for therapeutic intervention.
Pneumococci bind avidly to cells of the upper and lower respiratory tract. Like most bacteria, adherence of pneumococci to human cells is achieved by presentation of bacterial surface proteins that bind to eukaryotic cell surface carbohydrates in a lectin-like fashion (Cundell, D. and Tuomanen, E. (1994) Microb Pathog 17:361-374). Pneumococci bind to non-inflamed epithelium, a process that can be viewed as asymptomatic carriage. It has been proposed that the conversion to invasive disease involves the local generation of inflammatory factors which, activating the human epithelial cell, change the number and type of receptors available on the human cells (Cundell, D. et al. (1995) Nature, 377:435-438). Presented with an opportunity in this new setting, pneumococci appear to take advantage and engage one of these unregulated receptors, the platelet activating factor (PAF) receptor (Cundell et al. (1995) Nature, 377:435-438). Within minutes of the appearance of the PAF receptor, pneumococci undergo waves of enhanced adherence and invasion. Inhibition of bacterial binding to activated cells, for instance by soluble receptor analogs, blocks the progression to disease in animal models (Idanpaan-Heikkila, I. et al. (1997) J. Infect. Dis., 176:704-712). Particularly effective in this regard are soluble carbohydrates containing lacto-N-neotetraose with or without an additional sialic acid, which prevent pneumococcal attachment to human cells in vitro and prevent colonization in the lung in vivo.
Pneumococci display a family of surface proteins which bind to the bacterial surface by non-covalent association to the cell wall teichoic acid or lipoteichoic acid, specifically through its terminal component phosphorylcholine. The surface of Streptococcus pneumoniae is decorated with twelve types of these choline binding proteins (Cbps). The Cbps are represented by a family of molecules which decorate the surface of pneumococcus, each serving a unique function but bound to the surface by a common element. These proteins consist of an N-terminal activity domain and a repeated C-terminal signature choline binding domain that contains two to greater than ten repeats of a 20 amino acid choline binding sequence that binds to phosphoryl choline and that anchors these molecules to the surface of the bacteria. This motif has been identified in the C-terminal regions of a secreted glycoprotein from Clostridium acetobutylicum NCIB 88052 [Sanchez-Beato, et al., J. Bacteriol. 177:1098-1103 (1995)], toxins A and B from Clostridium difficile [Von Eichel-Streiber and Sauerbom, Gene 96:107-13 (1990); Von Eichel-Streiber et al., J. Bacteriol. 174:6707-6710 (1992)], a glucan-binding protein from Streptococcus mutans, several glycosyltransferases from Streptococcus downei and S. mutans, the murein hydrolase (LytA) from pneumococcus and pneumococcal lytic phage [Ronda et al., Eur. J. Biochem. 164:621-4 (1987); Diaz et al., J. Bacteriol. 174:5516-25 (1992); Romero et al., Microb. Lett. 108:87-92 (1993); Yother and White, J. Bacteriol. 176:2976-85 (1994)], and a surface antigen (PspA) also from pneumococcus. The known pneumococcal Cbp family members are CbpA, LytA and PspA.
The choline binding domain was recognized and fully characterized by Lopez et al. in his studies of the autolytic enzyme, LytA (Ronda et al. (1987) Eur. J. Biochem, 164:621-624). From studies of the sequences of the recognized choline binding proteins, consensus sequences have been reported, most particularly that of Garcia et al: GWLKDNGSWYYLNANGAMAT (SEQ ID NO:26) (Garcia, P. et al (1990) Gene 86:81-88; Wren B. et al (1991)Mol Microbiol 5:797-803); Sanchez-Beato, A. R. et al (1995) J Bacteriol 177:1098-1103).
Teichoic acid (TA), an integral part of the cell wall of Streptococcus pneumoniae, contains many terminal phosphorylcholine moieties. Choline affinity chromatography or Mono-Q Sepharose, a close relative of choline, were used to purify the CBPs. Previous studies have shown that PspA, as well as one other surface exposed protein, LytA, the autolytic amidase, bind in a choline-dependent manner. PspA, a protein having a molecular weight of 84 kDa, and which is highly variable, is released from the cell surface with high choline concentration (at least about 2% to about 10%). The function of PspA is unknown. LytA, or autolysin, is a 36 kDa protein, which lyses the pneumococcal cell wall (self lysis), but is not released from the cell by growth in high concentrations of choline, by washing in 10% choline, or by growth in ethanolamine. Reports on choline binding proteins include those by Sanchez-Puelles et al Gene 89:69-75 (1990), Briese and Hakenback Eur. J. Biochem. 146:417-427, Yother and White J. of Bacteriol. 176:2976-2985, Sanchez-Beato et al J. of Bacteriol. 177:1098-1103, and Wren Micro. Review Mol. Microbiol. 5:797-803 (1991), which are hereby incorporated by reference in their entirety.
CbpA is an adhesin (ligand) for the glycoconjugate containing receptors present on the surface of eucaryotic cells. CbpA is a 663 amino acid protein with an apparent molecular mass of 112 KDa. CbpA has been shown to be critical to pneumococcal colonization and attachment to human cells. Mutants with defects in cbpA show reduced virulence in the infant rat model for nasopharyngeal colonization and fail to bind to eucaryotic cells found at the site of infection and to glycoconjugates that bind pneumococcus. The CbpA protein cross reacts with human convalescent antisera, and antisera to the Cbps passively protected mice in a model for sepsis. Since the process of colonization and the progression to disease depend on pneumococcal attachment to human cells as a primary step, interruption of the function of surface choline binding proteins or the choline binding domain, for instance by cross reactive antibody or by inhibition with a competitive peptide mimicking this domain, may be relevant to blocking disease.
Choline-binding proteins for anti-pneumococcal vaccines are disclosed in International Patent Application WO 97/41151 of Masure, et al, which claims priority to U.S. patent application Ser. No. 08/642,250, which are hereby incorporated by reference in their entirety. The International patent Application WO 97/41151 discloses partial polypeptide sequences of certain putative choline binding proteins and the polypeptide sequence of CbpA.
The cell wall associated choline binding protein, LytA, a murein amidase, is an autolysin that is responsible for remodeling the cell wall, the separation of daughter cells, cell death in stationary phase and penicillin induced cell lysis. Expansion of the cell wall during bacterial growth and splitting of the septum for cell separation require enzymes that cleave the peptidoglycan network enclosing the cell. In addition to acting as spacemaker enzymes for cell wall growth, some of these enzymes act as autolysins, thereby representing potentially suicidal enzymes. Regulation of these enzymes is therefore important and must take into consideration their extracytoplasmic location. Unlike other bacteria which have multiple autolysins, S. pneumoniae has only one major autolysin, LytA. Antibiotics such as penicillin induce bacteriolysis by interfering with the control of the endogenous autolysins (Tomasz and Holtje, 1977, in Microbiology, D. Schlessinger, ed., pp. 202-215; Tomasz, 1983, in The Target of Penicillin, R. Hackenback et al eds., pp. 155-172). Although the binding of antibiotics to cell wall synthetic enzymes has been well characterized, the mechanism by which it induces autolysin mediated cell wall degradation is unknown. Tolerance to such antibiotics arises if the bacterial autolysins are not triggered as the antibiotic inhibits the cell wall synthetic machinery. The question of whether antibodies raised against LytA are protective against infection by S. pneumoniae and its requirement for virulence remains controversial. In some cases, loss of LytA by deletion leads to less virulence, while others report no effect (Berry, A. M. et al (1992) Microb Pathog 12:87-93).
PspA has been reported to be a candidate for a S. pneumoniae vaccine as it has been found in all pneumococci to date [see Yother et al., J. Bacteriol., 174:610 (1992)]; the purified protein can be used to elicit protective immunity in mice; and antibodies against the protein confer passive immunity in mice [Talkington et al., Microb. Pathog. 13:343-355 (1992)]. However, PspA demonstrates antigenic variability between strains in the N-terminal half of the protein, which contains the immunogenic and protection eliciting epitopes (Yother et al., supra). This protein does not represent a common antigen for all strains of S. pneumoniae, and therefore is not an optimal vaccine candidate.
As described above, each Cbp, while being bound to the surface by a common element, the choline binding domain, serves a unique function. This particular function is largely determined by the unique N-terminal activity domain of each Cbp. N-terminal domain fragments of Cpbs, particularly CbpA, have been shown to have activity in blocking adherence and are candidates for immunogenic vaccines, independent of the C-terminal choline binding domain. In particular, Tuomanen et al, U.S. Ser. No. 09/056,019, which is hereby incorporated herein by reference in its entirety, provides isolated polypeptides comprising N-terminal choline binding protein A truncates, particularly wherein the polypeptides do not bind to choline. Vaccines comprising such N-terminal truncates, DNA encoding such truncates, or antibodies directed against such truncates are also described in U.S. Ser. No. 09/056,019.
An additional surface exported protein that affects adherence and represents a virulence determinant for pneumococcus, ZmpB, has been identified and described by R. Novak and R. Masure in U.S. Ser. No. 09/096,336 filed Jun. 11, 1998 which is hereby incorporated by reference in its entirety. ZmpB is a zinc metalloprotease, dependent on Zn2+ for functional activity. zmpB mutants show loss of adherence and colonization and are defective in the autolytic pathway. A mutation in zmpB selectively alters the production of CbpA and generates covalently modified LytA. ZmpB is proposed to be a master regulatory protein that controls the expression of multiple Cbps critical for bacterial survival in the human host. ZmpB is a candidate for an S. pneumoniae vaccine or as one component of a multi-component vaccine.
The deficiencies of whole bacterial or capsule-based vaccines and the emerging problems with antibiotic resistance among infectious bacteria, including S. pneumoniae strains, have raised an urgent need for alternative effective vaccines and therapies. The invention herein fills such a need in providing protective vaccines and therapies.
The citation of references herein shall not be construed as an admission that such is prior art to the present invention.
In its broadest aspect, the present invention encompasses isolated polypeptides comprising an amino acid sequence of a choline binding protein, CbpG.
The present invention is directed to an isolated polypeptide comprising an amino acid sequence of a pneumococcal choline binding protein, CbpG and wherein such polypeptide contains a choline binding domain sequence which is homologous to SEQ ID NO:1.
The present invention provides an isolated polypeptide comprising an amino acid sequence of a pneumococcal choline binding protein, CbpG wherein such polypeptide contains a choline binding domain sequence which is homologous to GWLKDNGSWYYLNANGAMAT (SEQ ID NO:10).
The present invention further provides an isolated polypeptide comprising an amino acid sequence of an N-terminal CbpG choline binding protein truncate. The present invention provides an N-terminal CbpG truncate comprising the amino acid sequence as set forth in SEQ ID NO: 11. The invention particularly provides an N-terminal CbpG truncate consisting of amino acids 1-90 of the CbpG choline binding protein. In a further embodiment, the N-terminal CbpG truncate comprises less than amino acids 1-90 of the CbpG choline binding protein.
In a still further aspect, the present invention extends to vaccines based on the choline binding proteins described herein.
The present invention provides an isolated polypeptide comprising an amino acid sequence of a choline binding protein CbpG. The polypeptide comprises the amino acid sequence as set forth in SEQ ID NO:2 including fragments, mutants, variants, analogs, or derivatives, thereof The isolated polypeptide is suitable for use in immunizing animals and humans against bacterial infection, preferably pneumococci.
The present invention further provides an isolated polypeptide comprising an amino acid sequence of a N-terminal CbpG choline binding protein truncate, particularly wherein the polypeptide does not bind to choline. This invention provides an isolated immunogenic polypeptide comprising an amino acid sequence comprising an N-terminal truncate of a choline binding protein CbpG. In a particular embodiment, this invention provides an isolated immunogenic N-terminal CbpG truncate polypeptide comprising the amino acid sequence as set forth in SEQ ID NO:11. The present invention further particularly provides an immunogenic N-terminal CbpG truncate polypeptide consisting of amino acids 1-90 of CbpG, or of less than amino acid 1-90 of CbpG.
In a still further aspect, the present invention extends to an immunogenic choline binding protein polypeptide CbpG or a fragment thereof The present invention also extends to immunogenic choline binding protein polypeptides wherein such polypeptides comprise a combination of at least two choline binding polypeptides selected from the group consisting of CbpG and at least one other choline binding polypeptide, including fragments thereof and N-terminal truncates thereof.
The present invention also relates to isolated nucleic acids, such as recombinant DNA molecules or cloned genes, or degenerate variants thereof, mutants, analogs, or fragments thereof, which encode the isolated polypeptide or which competitively inhibit the activity of the polypeptide. The present invention further relates to isolated nucleic acids, such as recombinant DNA molecules or cloned genes, or degenerate variants thereof, mutants, analogs, or fragments thereof, which encode the choline binding protein CbpG. Preferably, the isolated nucleic acid, which includes degenerates, variants, mutants, analogs, or fragments thereof, has a sequence as set forth in SEQ ID NO:3. In a further embodiment of the invention, the full DNA sequence of the recombinant DNA molecule or cloned gene so determined may be operatively linked to an expression control sequence which may be introduced into an appropriate host. The invention accordingly extends to unicellular hosts transformed with the cloned gene or recombinant DNA molecule comprising a DNA sequence encoding the present invention, and more particularly, the DNA sequences or fragments thereof determined from the sequences set forth above.
In a particular embodiment, the nucleic acid has the sequence selected from the group comprising SEQ ID NO:3; a sequence complementary to SEQ ID NO:3; or a homologous sequence which is substantially similar to SEQ ID NO:3. In a further embodiment, the nucleic acid has the sequence consisting of SEQ ID NO:3.
The present invention further relates to isolated nucleic acids encoding an N-terminal choline binding protein truncate, particularly wherein the encoded polypeptide does not bind to choline. This invention particularly provides isolated nucleic acids encoding N-terminal choline binding protein truncates of choline binding protein CbpG. The invention further provides a nucleic acid encoding an N-terminal CbpG truncate comprising SEQ ID NO:12, a sequence complimentary to SEQ ID NO:12, or a homologous sequence which is substantially similar to SEQ ID NO:12. In a further embodiment, the nucleic acid encoding the N-terminal CbpG truncate has the sequence consisting of SEQ ID NO:12.
In a particular embodiment, the nucleic acid encoding an N-terminal CbpG truncate is capable of encoding amino acids 1-90 of CbpG. In a still further embodiment, the nucleic acid encoding an N-terminal CbpG truncate encodes a truncate smaller than amino acids 1-90 of CbpG.
Nucleic acid vaccines or DNA vaccines utilize nucleic acids encoding particular immunogenic polypeptides to induce immunity in a host against such encoded immunogenic vaccines. Such nucleic acid based vaccines can be used directly as naked DNA, or can utilize well recognized expression vectors or retroviral vectors, as more particularly described herein, to encode such immunogenic polypeptide on expression in the host cell. Methods to generate and utilize such any such nucleic acid vaccines or DNA vaccines are well known in the art.
The present invention relates to nucleic acid vaccines or DNA vaccines comprising nucleic acids encoding immunogenic polypeptides of a choline binding protein CbpG. The present invention relates to nucleic acid vaccines or DNA vaccines comprising nucleic acids encoding immunogenic polypeptides of choline binding protein CbpG or a fragment thereof or any combination of CbpG with at least one other choline binding polypeptide. Still further, this invention provides nucleic acid vaccines or DNA vaccines comprising nucleic acids encoding an immunogenic N-terminal polypeptide of choline binding protein CbpG.
Antibodies against the isolated polypeptide include naturally raised and recombinantly prepared antibodies. These may include both polyclonal and monoclonal antibodies prepared by known genetic techniques, as well as bi-specific (chimeric) antibodies, and antibodies including other functionalities suiting them for diagnostic use. Such antibodies can be used in immunoassays to diagnose infection with a particular strain or species of bacteria. The antibodies can also be used for passive immunization to treat an infection with Gram positive bacteria, particularly pneumococcus. These antibodies may also be suitable for modulating bacterial adherence including but not limited to acting as competitive agents.
It is still a further object of the present invention to provide a method for the treatment of mammals to control the amount or activity of the pneumococci bacteria or its subunits, so as to treat or avert the adverse consequences of invasive, spontaneous, or idiopathic pathological states. This invention provides pharmaceutical compositions for use in therapeutic methods which comprise or are based upon the isolated polypeptides, their subunits or their binding partners.
The invention further provides pharmaceutical compositions, vaccines, and diagnostic and therapeutic methods of use thereof
The invention provides pharmaceutical compositions comprising a choline binding polypeptide CbpG and a pharmaceutically acceptable carrier. The present invention further provides pharmaceutical compositions comprising a choline binding polypeptide of a choline binding protein CbpG, or a fragment thereof or any combination of CbpG with at least one other choline binding polypeptide, and a pharmaceutically acceptable carrier. Still further, this invention provides pharmaceutical compositions comprising N-terminal polypeptide of choline binding protein CbpG and a pharmaceutically acceptable carrier.
The invention further relates to a vaccine for protection of an animal subject from infection with a Gram positive bacterium comprising a vector containing a gene encoding a choline binding protein CbpG of a Gram positive bacterium operatively associated with a promoter capable of directing expression of the gene in the subject. Preferably, such vaccine contains a gene encoding a choline binding protein CbpG of pneumococcus.
In another aspect, the invention is directed to a vaccine for protection of an animal subject from infection with a Gram positive bacterium, most preferably pneumococcus, comprising an immunogenic amount of a choline binding protein CbpG, or a derivative or fragment thereof. Such a vaccine may contain the protein conjugated covalently to a bacterial capsule or capsules from one or more strains of bacteria, including pneumococcus. In one such embodiment, at least one of the bacterial capsules is derived from a mutant strain of bacteria which is non-adherent and non-virulent. In a further such embodiment, the non-adherent and non-virulent bacteria is a pneumococcus and is a CbpG mutant bacteria. Such non-adherent and non-virulent CbpG mutant bacteria can further be utilized in expressing other immunogenic or therapeutic proteins for the purposes of eliciting immune responses to any such other proteins in the context of vaccines and in other forms of therapy.
The invention further provides a cbpG mutant bacteria which is non-adherent to nasopharyngeal cells or to lung cells. Particularly, such cbpG mutant is a gram positive bacteria. More particularly, such cbpG mutant is Streptococcus. Most particularly, such cbpG mutant is Streptococcus pneumoniae.
The invention includes an assay system for screening of potential compounds effective to modulate the choline binding activity of the choline binding protein of the present invention. The invention more particularly includes an assay system for screening of potential compounds effective to modulate the choline binding activity of the choline binding protein CbpG. In one instance, the test compound, or an extract containing the compound, could be administered to a cellular sample expressing the choline binding protein CbpG to determine the compound""s effect upon the activity of the protein by comparison with a control. In a further instance the test compound, or an extract containing the compound, could be administered to a cellular sample expressing the choline binding protein CbpG, to determine the compound""s effect upon the activity of the protein, and thereby on adherence of said cellular sample to host cells, by comparison with a control.
The invention includes an assay system for screening of potential compounds effective to modulate the activity of the N-terminal domain of the choline binding protein of the present invention. The invention more particularly includes an assay system for screening of potential compounds effective to modulate the activity of the N-terminal domain of CbpG. In one instance, the test compound, or an extract containing the compound, could be administered to a cellular sample expressing the choline binding protein CbpG to determine the compound""s effect upon the activity of the protein by comparison with a control. In a further instance the test compound, or an extract containing the compound, could be administered to a cellular sample expressing the choline binding protein CbpG to determine the compound""s effect upon the activity of the protein, and thereby on adherence of said cellular sample to host cells, by comparison with a control. In another instance, the test compound, or an extract containing the compound, could be administered to a cellular sample expressing an N-terminal truncate (which lacks the C-terminal choline binding domain) of the choline binding protein CbpG to determine the compound""s effect upon the activity of the protein by comparison with a control. In a particular such instance, the cellular sample expresses an N-terminal CbpG truncate comprising amino acids 1-90 of CbpG or a smaller N-terminal truncate comprising less than amino acids 1-90 of CbpG. In a further such instance, the N-terminal CbpG truncate comprises the amino acid sequence set out in SEQ ID NO:11.
Other objects and advantages will become apparent to those skilled in the art from a review of the following description which proceeds with reference to the following illustrative drawings.

Identification and characterization of novel pneumococcal choline binding protein, CBPG, and diagnostic and therapeutic uses thereof


A blank for forming a carton, the carton formed therefrom, and a closure for the carton are provided for enclosing a plurality of unbound elongate smoking articles. The blank includes a base sub-blank at the distal end which is folded to form an open top box and a lid sub-blank at the proximal end which is folded to form a top for the box. A movable lid sub-panel is formed in the lid sub-blank which is hingedly removable therefrom to open the top of the box and which is reclosable by use of a foldable member provided therein. After opening of the movable lid sub-panel, the foldable member is folded over to form an extending tab which is received in a weakening line provided in an underlying front panel of the carton when it is desired to hold the movable lid sub-panel in a closed position.

Smoking articles such as cigarettes and various other articles are frequently provided in box shaped cartons or the like which can be opened in various manners. Typically, cigarettes are formed in packs of twenty, with ten packs then in turn being packaged in a paperboard carton in a 2×5 array. With such a carton, the packs themselves serve to seal and protect the enclosed cigarettes from the environment.
In order to reduce costs, it may be considered desirable to package cigarettes or the like in bulk in a carton with no other protection. Thus, there is a need for a carton which will protect the contents and which, once opened, will be reclosable to protect the contents.

Blank forming carton and closure therefor


Charged-particle-beam (CPB) exposure apparatus are disclosed that project a mask pattern onto a substrate while reducing aberrations. The mask pattern is typically subdivided into mask subfields, separated from each other by struts or the like, that are individually projected onto the substrate in an ordered manner to produce a die pattern in which constituent transfer subfields are properly stitched together. The apparatus comprises a projection-optical system, comprising multiple deflectors, between the mask and the substrate. The deflectors are connected to and controllably actuated by a controller in a manner serving to reduce third-order deflection aberrations to zero. Control of the deflectors is such that the charged-particle beam, propagating from the mask, has a trajectory that is a function of the location of the beam in a mask plane transverse to the optical axis and the location of the beam in a substrate plane transverse to the optical axis.

With the advent of microlithography methods and apparatus employing a charged-particle beam (CPB), a chronic difficulty has been achieving high exposure resolution and high throughput. Various approaches have been proposed to such end.
A first approach, tantalizingly offering prospects of high throughput, is an apparatus that exposes at least one entire die per exposure. One disadvantage of this approach is that it is difficult to manufacture a CPB-exposure mask from which an entire die can be projection-exposed in one shot. Another disadvantage is that it is exceedingly difficult to satisfactorily control aberrations over an entire die that is exposed in one shot. In fact, the goal of aberration control has been elusive with this first approach.
A second approach, offering prospects of improved resolution, utilizes "segmented exposure" in which a mask corresponding to an entire die is divided into multiple "subfields" that are individually exposed step-by-step. Thus, multiple shots are required to projection-expose a corresponding die on a sensitive substrate ("wafer"). Because the CPB is much smaller in transverse area using this second approach, the prospects for satisfactory aberration control (especially in peripheral regions of each shot) are more favorable.
An electron-optical theory applicable to certain types of segmented exposure is disclosed in Koikari et al., "Numerical Calculation On Optical System for EB Projection," Proceedings of the 9th International Microprocessing Conference ("Microprocess '96"), Jul. 8-11, 1996. Koikari et al. pertains to an electron-beam (EB) optical system employing a symmetric magnetic doublet lens system comprising first and second static axisymmetric lenses. The Koikari et al. CPB optical system also comprises multiple magnetic deflectors that impart paraxial deflections to an electron beam propagating, from a point on the mask, through the EB optical system to the wafer. The deflections purportedly reduce certain (but not all) aberrations. Also, the Koikari et al. deflectors impart no net deflection of the electron beam at the wafer plane. I.e., when electrons in the electron beam reach the wafer plane, all deflections imparted by the deflectors during propagation of the electrons from the mask to the wafer are exactly cancelled.
Consequently, the Koikari et al. theory, as presented in the Koikari et al. paper, can be applied only to segmented exposure in which individual mask subfields are contiguous (i.e., not separated from each other by boundary regions occupied by struts or the like).
Deflectors according to Koikari et al. are typically energized by respective power supplies under the control of a "controller" (i.e., numerical processor or computer). Each signal sent by the controller to a respective power supply is a function of the X- and Y-coordinates (X.sub.m, Y.sub.m) of the illuminated mask subfield but does not take into account the X- and Y-coordinates (X.sub.w, Y.sub.w) of the corresponding transfer subfield on the wafer.
FIG. 3 is a plan view of a wafer 11 exposed using a conventional electron-beam exposure system exhibiting aberration correction in a manner such as disclosed in Koikari et al. The optical axis AX of the system extends perpendicularly to the plane of the page at the position indicated. FIG. 3 also shows the relationship between the position of illuminated mask subfields 100a, 100b (relative to the optical axis AX) and the positions of the corresponding transfer subfields 200a, 200b (relative to the optical axis AX).
A CPB exposure apparatus embodying the principles of Koikari et al. has "zero deflection sensitivity" with respect to the deflectors, by which is meant that the deflectors collectively impart no net deflection of the electron beam from a wafer position that would be illuminated if no aberration-correcting deflectors were provided (or if the aberration-correcting deflectors were not energized. Also, according to conventional wisdom, the central coordinates (X.sub.w, Y.sub.w) of a transfer subfield are related to the central coordinates (X.sub.m, Y.sub.m) of the corresponding mask subfield by the following equations (1) and (2): EQU X.sub.w =-M(X.sub.m .multidot.cos .theta.-Y.sub.m .multidot.sin .theta.)(1) EQU Y.sub.w =-M(X.sub.m .multidot.sin .theta.+Y.sub.m .multidot.cos .theta.)(2)
wherein M is the combined magnification (M>0) of the static axisymmetric projection lenses and .theta. is a rotational angle of the image of the mask subfield formed on the respective transfer subfield.
To increase the mechanical rigidity of a segmented mask and render the mask more resistant to thermal distortion during illumination of the mask by the charged-particle beam, it is preferable to provide the mask with struts. Typically, the struts are placed between mask subfields. When the mask subfields on such a mask are projected onto the wafer, the struts are not projected. In order to form a complete integral image of the die on the wafer, all the transfer subfields of the die must be properly "stitched together." Thus, even though the mask subfields are separated from one another on the mask by the struts, the corresponding transfer subfields of the complete die formed on the wafer must be "connected" together without any intervening gaps. The charged-particle beam used to make such an exposure must be deflected in a manner ensuring that strut regions are not projected and that the transfer subfields in each die are properly stitched together on the wafer.
In order to stitch the transfer subfields together on the wafer when using a segmented mask with struts, aberrations must be controlled very well to ensure accurate alignment of all the transfer subfields with each other. Such aberrational control includes control of "hybrid" image-plane distortions and blur. (A "hybrid" distortion at a given point in an image is a function of both the distance of the point from the optical axis and the distance of the point from the center of the subfield. See Zhu et al., "Dynamic Correction of Aberrations in Focusing and Deflection Systems with Shaped Beams," SPIE 2522:66-77.) Functions according to, e.g., Koikari et al. are simply inadequate for use with a segmented mask including struts. Koikari et al. does not address how to make appropriate changes to the functions. Also, changing functions and/or adding more deflectors, without more, can undesirably cause aberrations to increase.
Therefore, there is a need for charged-particle-beam exposure apparatus that: (1) utilize a segmented mask including struts separating the mask subfields, (2) stitch together the corresponding transfer subfields on the wafer, and (3) satisfactorily reduce third-order aberrations.

Charged-particle-beam exposure apparatus exhibiting aberration control


An adjusting shaft for an electric regulating or control device, which operates equipment functions by means of a control cam, is profiled to engage the control cam and can be installed subsequent to the complete assembly of the control device and fixed relative to the control device by a spring detent and associated stop. The stop can also serve to prevent unintentional rotation of the control cam until displacement of the stop against the opposition of a biasing spring. The subsequently installed shaft can allow use of universal control devices with random shaft lengths and types.

Electric switches, regulators or control devices, such as those according to DE-C-26 04 783 (corresponding to GB 1 577 852) or DE-C-26 25 716 (corresponding to U.S. Pat. No. Re 31 597) have a driven element in the form of a cam disk on setting cam, which either directly or indirectly acts on contacts or switches an adjusting shaft, which drives the driven element; and locking elements driven by the adjusting shaft. The adjusting shafts are either fixed to the driven element or are inserted therein and consequently do not rotate relative to the driven element. The axial fixing of the shaft takes place by means of engagement in casing parts. This fixing can be brought about during the installation of the device by fitting an outer casing member.
As regulators or switches are often installed in devices that have widely differing designs positions of the operating or control elements, etc., it is necessary to provide different adjusting shaft lengths and shapes. It is also possible to connect additional or acessory switches to the regulators or switches other devices drive the switches by the same adjusting shaft. In this case, there must either be an indirect coupling, which operates less precisely due to its necessary clearance, or a separate switch or regulator type with a correspondingly lengthened spindle.

Adjustable device, particularly an electric switching, controlling or regulating device


Described are tray table assemblies that include an upper cover, a lower cover, and a support structure disposed between the upper cover and the lower cover. The support structure may include a plurality of upper protrusions extending toward an inner surface of the upper cover and a plurality of lower protrusions extending toward an inner surface of the lower cover such that the support structure provides structural rigidity to the tray table assembly. The plurality of upper protrusions may contact the inner surface of the upper cover, the plurality of lower protrusions may contact the inner surface of the lower cover, and the support structure may dissipate heat.

Many passenger seats such as those on passenger aircraft, buses, trains, and the like are arranged so that each passenger seat, other than the forward-most located passenger seats, faces the back of the next forward passenger seat. To increase a passenger's comfort, many passenger seat backs rotate between upright and reclined positions.
In some instances, a tray table may be mounted adjacent the back of each passenger seat for use by a passenger in the next aft passenger seat. The tray table is deployed by the passenger to provide a relatively flat surface for eating, working, recreation, or other uses.
In many conventional uses, the tray table may be mounted to the back of each passenger seat via a pair of retractable arms that allow the tray table to be pulled toward the passenger when deployed.
As shown in FIGS. 4A-4C, in a conventional configuration, a tray table 400 includes barrier layers as the upper 401 and lower 402 outer layers. Between the barrier layers 401 and 402, the tray table 400 also includes filler material 403, plastic supports 404, and a heat exchanger 405. The filler material may be, for example, an injected polyurethane foam material. There are numerous manufacturing concerns and problems related to foam including, for example, (1) significant scrap rates (leading to increases in manufacturing time and raw material cost), (2) extended manufacturing cycle times (due to curing, cooling, and inspecting the foam), (3) excess infrastructure and machine resources for both physical space and machine costs (e.g., foam manufacturing may require foam machines, ovens, cleaning stations, spray booth stations, mold racks, and molds), (4) increased human capital and labor costs, (5) high inspection failure rates due to delamination and/or warpage, (6) issues related to certification, and (7) health concerns.
To the extent the aforementioned problems are related to foam (i.e., filler material 403), these problems are eliminated by the current invention. The exploded assembly shown in FIG. 4C includes four components (filler material 403, two plastic supports 404, and a heat exchanger 405) that can all be replaced by a single support structure 105 (described in more detail below). In certain cases, replacing this group of components with support structure 105 results in (1) a raw material cost savings of more than $2 for every tray table assembly and/or (2) more than 20% reduction in weight compared to the group of components described above.
In certain situations, it may be desirable to eliminate filler material from tray tables to increase manufacturing efficiency and to reduce costs, while also providing a lightweight and durable tray table.


Directing an input light beam into a laser diode operated at a power below that required for lasing stimulates emissions, resulting in an amplified output beam. A dynamic steering system can focus the input beam onto front face of the laser diode. The steering system optionally includes an optical element mounted solely on piezoelectric actuators. Control signals for the actuators in the steering system control a base position and cause alternating movements of the optical element. A detector measuring optical power leaking from a back face of the laser diode can determine the power of the amplified beam that exits from the front face, and derivatives of the measured power with respect to the alternating movements indicate required adjustments of the base position. A polarizing beam splitter and quarter-wave plate in the path of beams can separate the input and amplified beams.

Heterodyne optical interferometers generally use input light beams that include two components of differing frequencies f1 and f2. One type of heterodyne interferometer uses one component (e.g., of frequency f1) in a measurement beam that is reflected from the object being measured and uses the other component (e.g., of frequency f2) in a reference beam. Analysis of the measurement and reference beams (after reflection) can determine a beat frequency that is equal to the frequency difference f1−f2 plus a Doppler shift Δf caused by reflection of the measurement beam from a moving object. The Doppler shift Δf, which may be small relative to the frequency f1 of the measurement beam, can be a significant fraction of the frequency difference f1−f2 and therefore can be precisely determined from the measured beat frequency. However, for an accurate indication of a Doppler shift Δf, the frequency difference f1−f2 in the input heterodyne beam should be greater than the maximum Doppler shift Δf encountered during measurements.
A Zeeman-split laser can provide an input light beam for a heterodyne interferometer. In a Zeeman-split laser, a magnetic field applied axially to a laser cavity creates a difference in the energies of emissions of left and right circularly polarized photons. As a result, the Zeeman-split laser has a gain curve for left circularly polarized light that differs from its gain curve for right circularly polarized light. If the laser cavity has an appropriate length, the Zeeman-split laser thus emits a beam including left circularly polarized light having one frequency and right circularly polarized light having another frequency. Unfortunately, the maximum frequency difference that can be practically created in a Zeeman-split laser is generally less than about 10 MHz, which may be smaller than the Doppler shift caused by reflection from a fast moving object.
An acousto-optic modulator (AOM) can be used with a laser (Zeeman-split or otherwise) to create or increase the frequency difference between the components beams. However, the robustness inherent of current AOM designs and required beam steering optics limit the usefulness of AOMs for creating high power heterodyne beams for precision interferometers.
A high power heterodyne input light beam is particularly important in an interferometer system where available space is limited but simultaneous measurements along many distinct axes are needed. In integrated circuit processing equipment, for example, a single heterodyne beam source may need to provide a heterodyne beam with sufficient power to simultaneously operate up to thirty or more interferometers. A beam source is thus needed that is capable of generating a heterodyne beam with a desired frequency difference and a power level suitable for multiple-axis interferometers.


A mounting bracket assembly for mounting a night vision device to a headmount apparatus, made up of an attachment member, a mounting bracket and a hinge. The attachment member attaches the mounting bracket assembly to the headmount. The mounting bracket is generally U-shaped for holding a night vision device. The mounting bracket includes a coupling pad disposed at an end of the mounting bracket for receiving a coupling member attached to the night vision device, and a spring clip attached to the coupling pad for locking the coupling member of the night vision device to the coupling pad. The hinge couples the mounting bracket to the attachment member, and includes a pivot mechanism for allowing articulated movement of the mounting bracket relative to the attachment member.

Night vision goggles, binoculars, and like devices are utilized in a variety of military and commercial applications where enhanced vision during low light or night conditions is desirable. Night vision devices typically utilize an image intensifier tube and associated optics for converting infrared and near infrared light reflected from objects, into viewable images. The United States Army currently uses a night vision goggle assembly referred to as the PVS-7B which is manufactured by ITT Electro-Optical Products Division (DEOP) of Roanoke, Va., the assignee herein. Also manufactured by ITT DEOP is a commercially available night vision binocular assembly referred to as the Night Quest.TM..
Night vision devices such as the PVS-7B night vision goggle assembly, are typically carried by a user by attaching the device to a headmount apparatus which is worn on the user's head. Such headmount apparatus typically consist of an adjustable headband/strap arrangement and a night vision device mounting bracket assembly. Night vision devices can also be carried by attaching them directly to a standard issue army helmet via a mounting bracket assembly which enables the night vision device to be selectively positioned in and out of the user's field of vision. An example of such a mounting bracket assembly is described in U.S. Pat. No. 4,449,787 to Burbo et al., which discloses a night vision imaging system mounted to a helmet via a helmet mount bracket assembly that is secured to the helmet at a front surface above the position of the eyes of a user. The mounting bracket assembly allows the night vision imaging system to be "flipped-up" away from the eyes of the user and locked in place when the night vision imaging system is not needed and "flipped-down" in front of the eyes of the user and locked in place when the night vision imaging system is needed.
A mounting bracket assembly for a PVS-7B night vision goggle assembly is described in U.S. Pat. No. 5,331,684 to Baril et al. The mounting bracket assembly described therein includes a support member for holding a PVS-7B or like night vision goggle. The support member is pivotally connected to a clamp mechanism which detachably couples the support member to a helmet. The mounting bracket assembly also includes a movable magnet and reed switch arrangement which activates and deactivates the night vision goggle assembly when it is "flipped" in and out of the user's field of vision.
The mounting bracket assemblies described above and in the prior art, do not generally allow the night vision device to be quickly removed from and/or installed thereon and are relatively heavy in weight. This is because these mounting bracket assemblies employ relatively complicated clamping and locking mechanisms for securing the night vision device to the mounting bracket assembly. The complexity of these clamping and locking mechanisms decreases the versatility of the night vision devices and the heavy weight of these mechanisms lead to increased user fatigue.
Accordingly there is a need for an improved night vision device mounting bracket assembly which allows quick removal and installation of the night vision device and has a reduced weight.

Mounting bracket assembly for a night vision device


The present invention, in certain aspects, discloses systems and methods for treating a human being and/or items with descenting material, the systems, in certain aspects, including a generator for producing descenting material, and, in certain aspects at least one direction apparatus in communication with the generator for receiving produced descenting material from the generator and for directing said descenting material in a desired direction.

1. Field of the Invention
The present invention is directed to descenting systems and methods; in certain aspects, to such systems and methods for facilitating human/animal encounters and activities; and in other aspects to methods and systems for reducing or eliminating human odors that are detectable by animals. The invention relates to a method of descenting the clothes and apparatus of sportsmen, both professional, non professional, military personnel, bikers, campers and the like. In certain aspects a method is provided for reducing or removing human scent and any other scent that is not advantageous in an environment from clothing and equipment of hunters and fish odors from fishermen utilizing an oxidizing agent which is ozone and/or a combination of hydroxyl and hydroperoxide ions. More particularly, there is provided a method of reducing or removing human scent and any other scent in a space between a human and an animal that is not advantageous in an environment, including scents emitting from human breath, human bodies, and from clothing and equipment, the methods utilizing an oxidizing agent which is, e.g., ozone and/or a combination of hydroxyl and hydroperoxide ions.
2. Description of Related Art
The prior art discloses a variety of descenting (removal of scents) apparatus and methods, examples of which (and not by way of limitation) are found in U.S. Pat. Nos. 4,309,388; 4,867,052; 4,941,270; 5,087,426; 5,433,919; 5,468,454; 5,484,472; 5,514,345; 5,539,930; 5,547,476; 5,667,564; 5,681,355; 5,762,648; 5,766,560; 5,789,368; 5,790,987; 5,911,957; 5,931,014; 6,007,770; 6,009,559; 6,134,806; 6,134,718; 6,149,038; 6,156,268; 6,163,098; 6,284,204; 6,312,507; 6,355,216; 6,379,435; 6,503,547; 6,564,591; 6,565,805; and 6,576,190, and published U.S. patent application 2003/0044308—all of which are incorporated fully herein for all purposes.
Hunters, wildlife enthusiasts, and wildlife photographers all have an interest in attracting wildlife, such as, but not limited to, deer; and/or an interest in not being detected by animals. Deer rely heavily on their sense of smell to react with their surrounding environment, including to sense danger, interact with other deer and find food. Scents that are not a natural part of the environment will often function as a warning to deer, which may result in the deer being spooked and running from the unnatural scent.
According to Bernier et al in Analytical Chemistry, 2000, volume 72, issue 4, pages 747-756 and references cited therein which are incorporated fully herein by reference, as many as 346 discernible compounds were identified in human skin emanations. The majority of these were carboxylic acids, alcohols and esters, but aldehydes, aromatics, heterocyclics, ketones, sulfides and thio compounds were also identified. Work cited in Bernier has identified over 100 compounds from human breath. Work cited in Bernier identified foot odor as another source of odor. Some of these compounds are the result of bacteria reacting with body emanations, while other compounds directly emanate from humans. Other compounds emanated from humans can include pheromones, deodorants and perfumes as well as the detergents, perfumes, scents, and additives left on human clothes. While it is not known which specific compound or blends of compounds emanating from humans are identified by an animal as human, there is currently no effective way to eliminate or reduce odors from humans and from clothing and equipment enough to reduce the odors to inhibit detection by wildlife or effectively attract wildlife.
Persons interested in preventing detection by deer detecting human odors or interested in attracting deer often use masks, attractants, or cover scents to prevent alerting and spooking the deer. Some commonly used masks are carbon sprays which, in addition to being dangerous to inhale and which can irritate skin, become ineffective once dry. Many of the attractants contain deer urine or estrous, which besides being offensive to the human user, have limited shelf life and are generally ineffective since especially the estrous tend to occur naturally only in certain seasons. Cover scents such as fruit extracts or fragrances last a short time and are often so over-powering that the deer easily identifies the smell as unnatural and runs. The use of descenting soaps and shampoos is messy, time-consuming, often skin irritating, often ineffective and does not address breath odor. Breath descenting using herbs are generally distasteful, and face masks containing carbons or sieves are extremely uncomfortable.
More recently, the use of clothing containing activated carbons and/or clothing containing bacteria killing metals such as silver has gained some popularity. However, activated carbon has a very low capacity for odorants and requires temperatures preferably above 400° C., more preferably above 600° C. to regenerate the carbon. These temperatures are well beyond temperature (100 C to 120 C) that a conventional gas or electric clothes dryer is capable of achieving. Placement of clothing in ovens capable of achieving 400 C plus temperatures needed to regenerate the carbon can damage the fabric of a garment. Silver or other metal-containing clothing requires direct contact of the metal with the bacteria to be effective, which is almost never the case since the clothing would then be so restricting as to be uncomfortable. These types of clothing are also expensive and do not address human odors such as those in human breath, nor do they address any of the odors emanating from the foot or any exposed part of the skin like the head and hands.
It has now been discovered that gaseous ozone effectively kills bacteria and reduces or eliminates odors emanating from humans as well as odors contained in clothing worn by hunters. The advantages of ozone over other known masking and descenting methods include the facts that: ozone is a gas that eliminates odors emanating from a person (e.g., a hunter) and from personal equipment and can eliminate odors in a space between a person and an animal; and ozone is completely natural to the environment and leaves behind a very pleasant clean smell that wildlife and humans readily recognize, e.g. after a lightning rain. Known ozone generators include electrical discharge, UV light, and combinations thereof. The generator may be battery operated, operated with a car adaptor, and/or may be operated with AC current. The AC current may be supplied directly from an electrical outlet, or may be supplied using a portable generator.
Ozone is well known to treat odorous air, microorganisms, bacteria, mold, smoke, aromatic hydrocarbons, and volatile organic compounds (see for example U.S. Pat. Nos. 1,961,878; 2,203,188; 3,421,836; 3,750,556; 3,937,967; 3,949,056; 4,045,316; 4,863,687; 4,904,289; 4,990,311; 5,087,426; 5,835,840; 5,983,834; 6,094,549; 6,613,277; 6,632,407; 20020030022; 20060096331; and references cited therein, which are all of which are incorporated fully herein for all purposes); and foreign references EP 261987; WO 200151096; WO 2003089017; WO 2005021135. WO 2005077425 and references cited therein, which are incorporated fully herein for all purposes, teaches the use of ozone to descent hunters clothing and other personal effects to be worn or carried on a hunting trip. In this case, the hunter, the clothing or personal effects are placed in a container, a portable enclosure, or a special descenting closet or room located, e.g., in a lodge or cabin in which the hunter is staying. As soon as the hunter leaves the building and enters a vehicle, or passes a moving vehicle, or begins to sweat, any prior descenting is of little value.
Animals have an acute sense of smell and are capable of recognizing a human scent or any other scent that is not advantageous in that environment at long distances. To avoid such recognition a hunter will attempt to stay down wind of the animal being hunted. Certain known methods used by hunters to trick animals are to mask the human odor utilizing a carbon spray or cover spray scents or an animal scent. Unfortunately the animal scents which are utilized, are obnoxious and linger on the clothing for long periods of time and often do not mask out human scents. Some of the scents utilized include animal urine. A hunter who is camping overnight does not desire the animal scents to be carried over to bedtime, home, car, etc.
There are other drawbacks in utilizing animal scents or any other scents. The scent may attract a predator of the game which the hunter is not hunting for which the hunter may not be prepared to encounter. Containers containing food, beverages, or any other substance emit scents readily recognizable to animals that may not be masked by animal scents or may not be natural to a given environment. Female hikers, campers, hunters, etc. can emit a readily recognizable scent to animals from menstruation that may not be masked by animal scents. Also, the weapon used by the hunter has an odor recognizable by some animals which cannot be disguised with a scent.
Fishermen have the problem of fish odor on their hands and clothes which is difficult to remove. For fishermen camping overnight the fish odor is not only undesirable because of the odor but can also attract animals such as bears which the fishermen is not prepared to meet.
Hunters have prepared their clothing before hand by washing to remove prior scents and/or human odor. The washing materials may also leave an odor. However, out in the field the hunter can sweat and permeate the clothing with a human scent. It would be desirable to deodorize clothing during a hunt or while on a fishing trip.
Ozone has been used for decontaminating buildings and for decolorizing denim garments. U.S. Pat. No. 5,833,740 to Brais discloses an apparatus for sterilizing bottles utilizing ozone. The reference recognizes that ozone in large quantities can be harmful or irritating. Consequently, it was necessary to provide means for decomposing the excess ozone and/or to cause its escape into the atmosphere.
Ozone is a powerful oxidizing agent. Ozone has 150% of the oxidizing potential of chlorine and twice the oxidizing potential of bromine. Ozone has been shown to be much more effective than chlorine with a reaction time up to 10 times faster. Ozone also readily self-destructs into simple diatomic oxygen due to its inherent instability. Ozone oxidizes biological products and kills bacteria.
Catalytic ionization of air using ultraviolet light is known to produce a mixture of ozone-containing hydroxyl and hydroperoxide ions. Ionization devices which are used to eliminate smoke and odors are known in the art to produce hydroxyl and hydroperoxide ions, e.g. those used in automobiles.


This invention relates to a method of producing purified methyl isobutyl ketone (MIBK) comprising subjecting a feed stream containing MIBK and impurities to a first distillation procedure from which acetone is recovered. The bottom product of the first distillation procedure is fed to a liquid-liquid separator and an organic phase from the said liquid-liquid separator is fed to the top region of a second distillation column to produce an overhead product. The said overhead product is condensed and fed to the said liquid-liquid separator. A bottom product containing MIBK is withdrawn from the second distillation column. This bottom product is fed to a third distillation column, high boiling impurities are withdrawn as a bottom product, and purified MIBK is also withdrawn. The invention also relates to an apparatus used in such a method.

Processes which include condensation of one or more carbonyl-containing reactants to form an unsaturated carbonyl-containing compound, and hydrogenation of said unsaturated compound to a saturated carbonyl-containing product, are well known.
One such process is the preparation of methyl isobutyl ketone (MIBK) from acetone. In this process condensation of two acetone molecules yield diacetone alcohol (DAA) which is dehydrated to yield mesityl oxide (MSO), and the MSO is hydrogenated to MIBK. The condensation and dehydration reaction takes place in the presence of an acidic catalyst, and the hydrogenation takes place in the presence of a hydrogenation catalyst such as a noble metal.
The production of MIBK can take place in two process steps as indicated above or in a single process step in the presence of a single condensation and hydrogenation catalyst. Such single step processes are disclosed in, for example, U.S. Pat. No. 3,574,763; EP 1 32 1 450 and South African complete patent application number 2004/8988.
The MIBK produced from the condensation and hydrogenation of acetone includes one or more impurities such as propane, isobutane, methyl pentane, acetone, 2-propanol, water, diacetone alcohol (DAA), mesityl oxide (MSO) and high boiling compounds such as diisobutyl ketone, C9 paraffins and ketones.
Methods of purifying MIBK are also known in the art. The article "Why not do it in one step", Chemtech, January 1977 discloses a process wherein the MIBK reaction products are subjected to a gas separator to remove unreacted hydrogen for recycle to the MIBK reactor. The liquid from the gas separator is fed to a first distillation column where acetone is recovered as an overhead product and recycled to the MIBK reactor. The bottom product of the first column is then fed to a liquid-liquid separator (decanter), where the aqueous phase is removed from the process and the organic phase is fed to a second distillation column somewhere between the take-off of the bottom product and a reflux entry position. An overhead low boiling product is removed as distillate in the second column. The bottom product of the second column is then fed to a third distillation column where high boiling compounds are removed as the bottom product and MIBK is removed as the distillate.
"Methyl Isobutyl Ketone by Direct Condensation of Acetone", SRI Reports, May 1972, discloses a similar process as described above. The main difference is that the first distillation column described above is replaced by two columns, namely a first distillation column where low boiling products (particularly methyl pentane as an azeotrope with acetone) are removed as a distillate. The bottom product is then fed to a second column where the unreacted acetone is removed as a distillate and recycled to the MIBK reactor. Another difference is that the third column (similar function as the second column above) is fitted with an overheads decanter.

Method and apparatus for producing purified methyl isobutyl ketone


The invention relates to unsymmetrical cyanine dyes with a cationic side chain, typically benzothiazole or benzoxazole derivatives, that exhibit enhanced fluorescence on binding with DNA or RNA, where such fluorescence can be used for evaluating the presence of nucleic acid polymers. The dyes generally have the formula: ##STR1## where R.sup.1 is an alkyl group having 1-6 carbons; X is 0, S, or NR.sup.2, where R.sup.2 is an alkyl group having 1-6 carbons, or CR.sup.3 R.sup.4, where R.sup.3 and R.sup.4, which may be the same or different, are alkyl groups having 1-6 carbons;. n=0, 1, or 2; PA0 Z.sup.1 and Z.sup.2, which may be the same or different, are independently hydrogen, an alkyl group having 1-6 carbons, or aryl, or Z.sup.1 and Z.sup.2 taken in combination complete a 6-membered aromatic ring; PA0 Y is HC.dbd.CH; and PA0 each of P and M=0 or 2, such that P+M=1; and PA0 TAIL is an aminoalkyl chain containing a backbone of 3-42 carbons and 1-5 positively charged nitrogens intermittently or equally spaced within the backbone, such that there are at least two carbons between sequential nitrogens.

Fluorescent dyes have many uses and are known to be particularly suitable for biological applications in which the high detectability of fluorescence is desirable. By binding to a specific biological ingredient in a sample, a fluorescent dye can be used to indicate the presence or the quantity of the specific ingredient in a sample. A variety of fluorescent dyes are available for specific fluorescent staining and quantitation of DNA and RNA, and other applications involving nucleic acids.
Unsymmetrical cyanine dyes were described long before much was known about DNA, by Brooker, et al., J. AM. CHEM. SOC. 64, 199 (1942). Some dyes in this class have since been found to be useful in fluorescent staining of DNA and RNA. The dye sold under the name Thiazole Orange has particular advantages in the quantitative analysis of immature blood cells or reticulocytes. U.S. Pat. No. 4,883,867 to Lee, et al. (1989) ('867 patent); Lee, et al., Thiazole Orange: A New Dye for Reticulocyte Analysis, CYTOMETRY 7, 508 (1986). As indicated in the '867 patent to Lee, et al., the dye used for this purpose must be able to penetrate the cell membrane.
The inventors have discovered that a composition that includes a side chain with additional positive charges is a polar compound that is unable to readily penetrate cell membranes. Nevertheless, the dye discovered by inventors is highly useful as a stain for nucleic acids because it has an extremely high affinity for nucleic acid polymers that is greater than that of the parent compounds (e.g. thiazole orange, oxazole yellow, thiazole red). The dyes of the invention are sensitive to even small quantities of nucleic acid polymers not contained inside living cells, e.g. in cell extracts, as well as to nucleic acids in permeabilized cells, such as dead or fixed cells. The combination of dye with nucleic acid polymers generally results in fluorescence that is linearly related to the amount of polymer present in the sample.
The novel fluorescent dyes is similar to another discovery of inventors, DIMERS OF UNSYMMETRICAL CYANINE DYES (Ser. No. 07/761,177 filed Sep. 16, 1991 by Yue, Johnson & Haugland). The dimer compounds contain two unsymmetrical cyanine dye units linked by a bridge having one or more positive charges, whereas the monomer dyes of the instant invention contain only one unsymmetrical cyanine dye unit. The affinity of the dimer for nucleic acid polymers is even greater than that of the subject invention. For some applications high affinity may cause problems where removal of the dye to allow recovery of the nucleic acid polymers is desired. It is advantageous to have a dye that has an affinity for nucleic acids that is not so strong as to require harsh conditions to separate the bound dye from the nucleic acid, yet is strong enough to permit rapid binding to and quantitative detection of the polymer, with minimum background from unbound dye. In addition, the monomer dye with cationic side chain unexpectedly shows less optical deterioration in aqueous solutions than the dimer. This characteristic of the monomer dyes in aqueous solutions is advantageous in that the dye continues to be useful in the evaluation of nucleic acids even under prolonged experimental conditions.

Unsymmetrical cyanine dyes with a cationic side chain


A method is provided in one example embodiment executed at a service metering framework (SMF) engine including a processor, and includes interfacing, by an event listener at the SMF engine, with an application being executed in a cloud by a remote client device, detecting a metering event associated with the application during execution of the application, receiving a value of at least one metering attribute associated with the metering event, and storing the at least one metering attribute and the value as a formatted metered record in a SMF database searchable according to the metering attribute. In a specific embodiment, the event listener exposes an application programming interface (API) of the SMF engine to the application to facilitate definitions of the metering event and the at least one metering attribute in the application.

Cloud computing represents a paradigm shift that benefits individual consumers, enterprises, and service providers. Information technology (IT) resources and services in the cloud may be abstracted from underlying infrastructure and provided on demand and at scale in a shared multitenant and elastic environment. Cloud services may be characterized by various parameters, including a payment or pricing model (e.g., pay-as-you-go, usage based pricing), elasticity (e.g., users can dynamically consume more or less resources), location independence, high fault tolerance, high availability, and ubiquitous access to services (e.g., users can access the services from any location, using any device). Cloud services include storage, network, and computing, facilitating various service models, such as infrastructure as a service (IaaS), platform as a service (PaaS) and software as a service (SaaS). The increasing number of applications, data and services offered in the cloud pose a challenge to service providers and enterprises in terms of various parameters, including managing the demand, and metering the usage.

System and method for a service metering framework in a network environment


To improve the efficiency of a reduction reaction. A power source applies a voltage to an oxidation electrode immersed in an aqueous solution in an oxidation tank and a reduction electrode immersed in an aqueous solution in a reduction tank, the voltage having a voltage value that changes with a predetermined cycle to be a voltage value at which ions can be desorbed from a surface of the oxidation electrode and a surface of the reduction electrode during one cycle of the voltage change. The frequency of the voltage is set within a range of 10 Hz to 1 kHz.

A conventional electrolytic reduction device is configured such that an oxidation tank in which an oxidation electrode is immersed in an aqueous solution and a reduction tank in which a reduction electrode is immersed in an aqueous solution are connected via a proton exchange membrane, and carbon dioxide is constantly supplied to the reduction tank via a tube. By connecting a power source between the electrodes and applying a constant DC voltage which has a value high enough for reactions of the oxidation electrode and the reduction electrode to proceed, protons and carbon dioxides are reduced on the reduction electrode side to produce hydrogen, carbon oxide, methane, ethylene, methanol, ethanol, and formic acid.

Electrolytic reduction device and electrolytic reduction method

NIPPON TELEGRAPH AND TELEPHONE CORPORATION


A transportable container for electrical devices having aluminum bottom, top and end panels. The top and bottom panels are configured having spaced apart interior and exterior sheet members laser welded to a corrugate stiffener. Each end panel is structured with an exterior sheet member spaced from an interior plurality of drawer guides laser welded to a corrugate stiffener. A plurality of aluminum drawers are mounted upon the drawer guides, each having an air entrance opening at its back communicating with air transfer outlets at a fan charged air distribution plenum. Conditioned a. c. power is provided at receptacles within each drawer as well as LAN connectors.

As systems such as aircraft both commercial and military have evolved, their technical complexity has advanced exponentially. In concert with such technical advancement the logistics associated with their maintenance, upgrading and repair has required the development of extensive information which traditionally has been generated in the form of paper carried printed text and drawings. The bulk of that paper text, sometimes referred to as "technical orders" (TO) can be substantial, paper technical orders for a complex military aircraft typically requiring palletizing for its transport. As a consequence of this information growth, industry and government have or are in the process of converting this extensive paper carried information into an electronic data collection manifested in technician carried laptop computers, personal data assistance (PDAs) and the like.
Accordingly, logistic concerns now have turned to the management of an extensive number of these portable electronic data devices. For instance, each one of a modern military fighter aircraft typically will require about three laptop computers with associated battery chargers and local area network (LAN) coupling capabilities for receiving TO upgrades and downloading maintenance data. Such logistical support components will be co-deployed with an extra base deployment of their corresponding aircraft. Often at a given base to which the aircraft are assigned, a collection of many laptop computers will be observed on the tops of tables connected with battery chargers, connected in turn to a. c. strip outlets, LANs and the like, a logistic management approach deemed less than desirable. Deployment of this equipment, for instance, to foreign bases requires some form of containment for shipping. Fiberglass transit cases capable of being lifted by two to four men have been available for this purposes. Suggestion has been made to retrofit these hard tooled cases with metal shelving for carrying the equipment. The result of such retrofitting is an excessive case weight and a support approach exhibiting little system design flexibility. Shelf containing steel cases for carrying equipment have been marketed. However, such containment also exhibits excessive weight.

Transportable container for electrical devices