Invaplex from gram negative bacteria, method of purification and methods of use

Abstract

Invaplex, a novel composition comprising invasin proteins and LPS from gram-negative bacteria is described as well as methods of using the novel composition as an adjuvant and a diagnostic tool.

1 . A composition comprising an isolated Invaplex of gram-negative bacteria comprising at least one invasin protein associated with LPS of said gram-negative bacteria.  
     
     
         2 . A composition according to  claim 1  wherein, said gram-negative bacteria is selected from the genera consisting of  Shigella, Escherichia, Salmonella, Yersinia, Rickettsia, Brucella, Erhlichiae, Edwardsiella, Campylobacter, Legionella  and  Neisseria.    
     
     
         3 . A composition according to  claim 2  wherein, said  Shigella  is selected from the group consisting of  S. flexneri, S. sonnei, S. boydii, S. dyesenteraie.    
     
     
         4 . A composition according to  claim 2  wherein, said  Escherichia  is  Escherichia coli.    
     
     
         5 . A composition according to  claim 4  wherein, said  Escherichia coli  is EIEC.  
     
     
         6 . A composition according to  claim 3  wherein said Invaplex comprises at least one invasin protein chosen from the group consisting of IpaA, IpaB, IpaC, IpaD, and LPS.  
     
     
         7 . A composition according to  claim 7  wherein said Invaplex comprises Ipaa, IpaB, IpaC, IpaD, and LPS.  
     
     
         8 . A composition according to  claim 7  wherein said Invaplex further comprises VirG or portions thereof.  
     
     
         9 . A composition according to  claim 5  wherein said Invaplex comprises at least one invasin protein chosen from the group consisting of IpaA, IpaB, IpaC, IpaD and LPS.  
     
     
         10 . A composition according to  claim 5  wherein said Invaplex comprises IpaA, IpaB, IpaC, and LPS.  
     
     
         11 . A composition according to  claim 10  wherein said Invaplex further comprises IpaD.  
     
     
         12 . A composition according to  claim 11  wherein said Invaplex further comprises VirG or portions thereof.  
     
     
         13 . A method for preparing isolated Invaplex from  Shigella,  said method comprising the steps of: 
 (i) water extracting  Shigella;      (ii) separating and discarding membrane fragments from said water extracted  Shigella  resulting in a solution containing the Invaplex; and    (iii) isolating the Invaplex from said solution.    
     
     
         14 . The method according to  claim 13  wherein said isolating in step iii is by using an ion exchange matrix.  
     
     
         15 . A method for preparing isolated Invaplex from  Escherichia,  said method comprising the steps of: 
 (i) water extracting  Escherichia;      (ii) separating and discarding membrane fragments from said water extracted  Escherichia  resulting in a solution containing the Invaplex; and    (iii) isolating the Invaplex from said solution.    
     
     
         16 . The method according to  claim 15  wherein said isolating in step iii is by using an ion-exchange matrix.  
     
     
         17 . A method for screening agents or drugs which reduce or eliminate Invaplex virulence said method comprising detecting a dissociation of said Invaplex in the presence of said agent or drug.  
     
     
         18 . A method for detecting gram-negative bacterial infection in a biological sample comprising 
 (i) contacting a sample with a solid surface to which is attached an Invaplex from bacteria suspected of causing the bacterial infection; and    (ii) detecting the presence or absence of a complex formed between said Invaplex and antibodies specific therefor in said sample wherein the presence of a complex formed indicates the presence of said bacterial infection.    
     
     
         19 . The method of  claim 18  wherein said biological sample is from an animal.  
     
     
         20 . A method for detecting gram-negative bacteria infection in a sample comprising (i) contacting a sample with a solid surface to which is attached an Invaplex from bacteria suspected of being present in said sample; 
 (ii) detecting the presence or absence of a complex formed between said Invaplex and said bacteria in said sample wherein the presence of a complex formed indicates the presence of said bacteria.    
     
     
         21 . The method of  claim 20  wherein said sample is chosen from the group consisting of: an environmental sample, an insect, and an animal.  
     
     
         22 . An antibody to an Invaplex which recognizes the Invaplex or any portion thereof.  
     
     
         23 . A method to elicit an antigen-specific immune response in a subject, said method comprising administering to said subject an Invaplex from a gram-negative bacteria along with said antigen wherein said antigen-specific immune response is chosen from the group consisting of cell-mediated immune response, humoral immune response, and mucosal immune response.  
     
     
         24 . The method of  claim 23  wherein said antigen is selected from the group consisting of viral antigens, mammalian cell surface molecules, bacterial antigens, fungal antigens, protozoan antigens, parasitic antigens, and cancer antigens.  
     
     
         25 . The method of  claim 23  wherein said Invaplex is administered by a route selected from the group consisting of intramuscular, bronchial, genital, nasal, oral, parenteral, transcutaneous, transdermal and rectal.