Method and materials for the identification of lipopolysaccharide producing microorganisms

Claim 2

An immunological reagent for the detection of two more endotoxin lipopolysaccharide/glycolipid antigen producing microorganisms comprising a monoclonal antibody specifically reactive with shared antigenic determinatnts of said lipopolysaccharide/glycolipid antigen said determinants being selected from the group consisting of the Lipid A region and core oligosaccharide region of said lipopolysaccharide/glycolipid antigen and said antibody being produced by a hybridoma selected from the group consisting of clones Re-1 (ATCC No. HB 8482), Re-2 (ATCC No. HB 8481), LA-4, (ATCC No. HB 8480), LA-5 (ATCC No. HB 8479) and C-2 (ATCC No. HB 8478).

Claim 5

The method according to claim 2 wherein said antibody is analytically detected by fluorometric, enzymatic or radiometric measurement of fluoroscent, enzymatic or radioactive material, chemically complexed with said antibody.

Claim 6

The method according to claim 2 wherein said indicatable antibody is analytically indicated by reaction with a second antibody specific for said indicatable antibody, said reaction being indicated by fluorometric, enzymatic or radiometric measurement of fluoroscent, enzymatic or radioactive material chemically complexed to either of said antibodies.

Claim 7

The method according to claim 2 wherein said region is the Lipid A region.

Claim 8

The method according to claim 2 wherein said region is the core-oligosaccharide region.

Claim 9

The method according to claim 2 wherein said sample is a clinical sample selected from the group consisting of blood, serum, saliva, stool, urine, milk, topical washing of skin or genitals, tissue samples or homogenates thereof and samples of cell culture fluids of infected cells or homogenates thereof.

Claim 10

In a two-site immunometric assay to identify an antigen in a sample comprising forming a ternary complex of a first labelled antibody, said antigen, and a second antibody said second antibody being bound to a solid carrier insoluble under ternary complex forming conditions wherein the presence of the antigen is detected and identified by measuring either the amount of labelled antibody bound to said solid carrier or the amount of unreacted labelled antibody: the improvement comprising employing as said second antibody a monoclonal antibody characterized in having affinity for an antigenic determinant site of said antigen which is shared by a plurality of taxonomically distinct microorganisms said determinant being selected from the group consisting of the Lipid A region and core oligosaccharide region of an antigen assocated with endotoxin molecules and said antibody being produced by a hybridoma selected from the group consisting of clones Re-1 (ATCC No. HB 8482), Re-2 (ATCC No. HB 8481), LA-4, (ATCC No. HB 8480), LA-5 (ATCC No. HB 8479) and C-2 (ATCC No. HB 8478).

Claim 11

In a two-site immunometric assay to identify an antigen in a sample comprising forming a ternary complex of a first labelled antibody, said antigen, and a second antibody said second antibody being bound to a solid carrier insoluble under ternary complex forming conditions wherein the presence of the antigen is detected and identified by measuring either the amount of labelled antibody bound to said solid carrier or the amount of unreacted labelled antibody: the improvement comprising employing as said second antibody a monoclonal antibody characterized in having affinity for an antigenic determinant site of said antigen which is shared by a plurality of taxonomically distinct microorganisms said determinant being selected from the group consisting of the Lipid A region and core oligosaccharide region of an antigen assocated with endotoxin molecules and said antibody being produced by a hybridoma selected from the group consisting of clones Re-1 (ATCC No. HB 8482), Re-2 (ATCC No. HB 8481), LA-4, (ATCC No. HB 8480), LA-5 (ATCC No. HB 8479) and C-2 (ATCC No. HB 8478).

Claim 12

The method according to claim 8 wherein said region is the Lipid-A region.

Claim 13

The method according to claim 8 wherein said region is the core oligosaccharide.

Claim 14

The method according to claim 8 wherein said antigenic determinant site reactive with said first antibody and said antigenic determinant site reaction with said second antibody are regions of the same macromolecule.

Claim 15

The method according to claim 11 wherein said antigenic determinant site reaction with said first antibody is comprised of the somatic region of an endotoxic lipopolysaccharide.

Claim 16

The method according to claim 11 wherein said antigenic determinant site reactive with said second antibody and said antigenic determinant site reactive with said first antibody are regions of separate macromolecules.

Claim 17

The method according to claim 13 wherein said antigenic determinant site reactive with said second antibody is the Lipid A or core oligosaccharide region of a lipopolysaccharide/glycolipid antigen associated with endotoxin molecules.

Claim 18

The method according to claim 13 wherein said antigenic determinant site reactive with said first antibody is a region of any nonendotoxic lipopolysacchride molecule with the proviso that said determinant is specific to a taxonomically distinct microorganism.

Claim 19

The method according to claim 11 wherein said first labelled antibody is labelled with a fluorometric, enzymatic or radioactive material.

Claim 20

A monoclonal antibody produced by a hybridoma formed by a fusion of cess from a mouse myeloma and spleen cells from a mouse previously immunized with Salmonella Re endotoxin glycolipid whereins said hybridoma is selected from the group consisting of Clone Re-1 (ATCC No. HB 8482) and Re-2 (ATCC No. 8481).

Claim 21

A monoclonal antibody produced by a hybridoma formed by a fusion of cess from a mouse myeloma and spleen cells from a mouse previously immunized with Salmonella Re endotoxin glycolipid whereins said hybridoma is selected from the group consisting of Clone Re-1 (ATCC No. HB 8482) and Re-2 (ATCC No. 8481).

Claim 22

A monoclonal antibody produced by a hybridoma, formed by a fusion of cells from a mouse myeloma and spleen cells from a mouse previously immunized with Lipid A wherein said hybridoma is selected from the group consisting of Clon LA-4 (ATCC No. HB 8480) and LA-5 (ATCC No. HB 8479).

Claim 23

A monoclonal antibody produced by a hybridoma, formed by a fusion of cells from a mouse myeloma and spleen cells from a mouse previously immunized with Lipid A wherein said hybridoma is selected from the group consisting of Clon LA-4 (ATCC No. HB 8480) and LA-5 (ATCC No. HB 8479).

Claim 24

The monoclonal antibody produced by the hybridoma is Clone C-2 (ATCC. No. HB 8478).

Claim 25

The monoclonal antibody produced by the hybridoma is Clone C-2 (ATCC. No. HB 8478).

Claim 26

A kit for use in the immunological detection of an antigen shared by a plurality of taxonomically distinct microorganisms comprising antigen reactive means comprising a monoclonal antibody specifically reactive with said antigen, said antigen possissing antigenic determinants selected from the group consisting of the Lipid A region and core oligosaccharide region of an antigen associated with endotoxin molecules, said antibody being further characterized in being chemically complexed with an analytically detectable reagent and being produced by a hybridoma selected from the group consisting of clones Re-1 (ATCC No. HB 8482), Re-2 (ATCC No. HB 8481), LA-4, (ATCC No. HB 8480), LA-5 (ATCC No. HB 8479) and C-2 (ATCC No. HB 8478).

Claim 27

A kit for use in the immunological detection of an antigen shared by a plurality of taxonomically distinct microorganisms comprising antigen reactive means comprising a monoclonal antibody specifically reactive with said antigen, said antigen possissing antigenic determinants selected from the group consisting of the Lipid A region and core oligosaccharide region of an antigen associated with endotoxin molecules, said antibody being further characterized in being chemically complexed with an analytically detectable reagent and being produced by a hybridoma selected from the group consisting of clones Re-1 (ATCC No. HB 8482), Re-2 (ATCC No. HB 8481), LA-4, (ATCC No. HB 8480), LA-5 (ATCC No. HB 8479) and C-2 (ATCC No. HB 8478).

Claim 28

The kit of claim 20 wherein said reagent comprises fluorescent, enzymic or radioactive material.

Claim 29

The kit of claim 21 wherein said reagent is the enzyme alkaline phosphatase.

Claim 30

The kit of claim 20 wherein said antibody is the monoclonal antibody produced by a hybridoma selected from the group consisting of Clone Re-1 (ATCC No. HB 8482) and Re-2 (ATCC No. HB 8481).

Claim 31

The kit of claim 20 where said antibody is the monoclonal antibody produced by a hybridoma selected from the group consisting of Clone LA-4 (ATCC No. HB 8480) and LA-5 (ATCC No. HB 8479).

Claim 32

The kit of claim 20 wherein said antibody is the monoclonal antibody produced by the hybridoma Clone C-2 (ATCC No. HB-8478).

Claim 33

A kit for use in the immunological detection of two or more lipopolysaccharide/glycolipid antigen associated with endotoxin molecules producing microorganisms comprising: (1) a lipopolysaccharide reactive means comprising a first monoclonal antibody specifically reactive with, an antigenic determinant selected from the group consisting of the Lipid A region and core oligosaccharide region of an antigen associated with endotoxin molecules, said antibody being produced by a hybridoma selected from the group consisting of clones Re-1 (ATCC No. HB 8482), Be-2 (ATCC No. HB 8481), LA-4, (ATCC No. HB 8480), LA-5 (ATCC No. HB 8479) and C-2 (ATCC No. HB 8478) and (2) a second antibody specifically reactive with said first antibody, said second antibody being further characterized in having chemically complexed thereto an analytically detectable reagent.

Claim 34

A kit for use in the immunological detection of two or more lipopolysaccharide/glycolipid antigen associated with endotoxin molecules producing microorganisms comprising: (1) a lipopolysaccharide reactive means comprising a first monoclonal antibody specifically reactive with, an antigenic determinant selected from the group consisting of the Lipid A region and core oligosaccharide region of an antigen associated with endotoxin molecules, said antibody being produced by a hybridoma selected from the group consisting of clones Re-1 (ATCC No. HB 8482), Be-2 (ATCC No. HB 8481), LA-4, (ATCC No. HB 8480), LA-5 (ATCC No. HB 8479) and C-2 (ATCC No. HB 8478) and (2) a second antibody specifically reactive with said first antibody, said second antibody being further characterized in having chemically complexed thereto an analytically detectable reagent.

Claim 35

The kit of claim 26 wherein said reagent comprises fluoroscent, enzymic, or radioactive material.

Claim 36

The kit of claim 27 wherein said reagent is the enzymic alkaline phosphatase.

Claim 37

The kit of claim 26 wherein said first antibody is the monoclonal antibody produced by a hybridoma selected from the group consisting of Clone Re-1 (ATCC No. HB 8482) and Re-2 (ATCC No. HB 8481).

Claim 38

The kit of claim 26 wherein said first antibody is the monoclonal antibody produced by a hybridoma selected from the group consisting of Clone LA-4 (ATCC No. HB 8480) and LA-5 (ATCC No. HB 8479).

Claim 39

The kit of claim 26 wherein said first antibody is the monoclonal antibody produced by the hybridoma Clone C-2 (ATCC No. HB-8478).

Claim 40

A kit for use in a two-site immunometric assay for the identification of a plurality of taxonomically distinct lipopolysaccharide producing microorganisms comprising: (1) a first lipopolysaccharide reactive means comprising a first antibody specifically reactive with an antigenic determinant of said lipopolysaccharide said determinant being specific to a taxonomically distinct microorganism, said first antibody being further characterized in having chemiclaly complexed thereto an analytically detectable reagent, and (2) a second lipopolysaccharide detecting means comprising a second monoclonal antibody specifically reactive to an antigenic determinant of lipopolysaccharide, selected from the group consisting of the Lipid A region and core oligosaccharide region, said antibody being produced by a hybridoma selected from the group consisting of clones Re-1 (ATCC No. HB 8482), Re-2 (ATCC NO. HB 8481), LA-4, (ATCC No. HB 8480), LA-5 (ATCC No. HB 8479) and C-2 (ATCC No. HB 8478).

Claim 41

A kit for use in a two-site immunometric assay for the identification of a plurality of taxonomically distinct lipopolysaccharide producing microorganisms comprising: (1) a first lipopolysaccharide reactive means comprising a first antibody specifically reactive with an antigenic determinant of said lipopolysaccharide said determinant being specific to a taxonomically distinct microorganism, said first antibody being further characterized in having chemiclaly complexed thereto an analytically detectable reagent, and (2) a second lipopolysaccharide detecting means comprising a second monoclonal antibody specifically reactive to an antigenic determinant of lipopolysaccharide, selected from the group consisting of the Lipid A region and core oligosaccharide region, said antibody being produced by a hybridoma selected from the group consisting of clones Re-1 (ATCC No. HB 8482), Re-2 (ATCC NO. HB 8481), LA-4, (ATCC No. HB 8480), LA-5 (ATCC No. HB 8479) and C-2 (ATCC No. HB 8478).

Claim 42

The kit of claim 32 wherein said reagent comprises fluoroscent, enzymic, or radioactive material.

Claim 43

The kit of claim 33 wherein said reagent is the the enzyme alkaline phosphataste.

Claim 44

The kit of claim 32 wherein said microorganisms are selected from the group consisting of Escherichia, Salmonella, Brucella, Neiserria, Chlamydia, and Rickettsia.

Claim 45

The kit of claim 32 wherein said second antibody is the monoclonal antibody produced by a hybridoma selected from the group consisting of Clone Re-1 (ATCC No. HB 8482) and Re-2 (ATCC No. HB 8481).

Claim 46

The kit of claim 32 wherein said second antibody is the monoclonal antibody produced by a hybridoma selected from the group consisting of Clone LA-4 (ATCC No. HB 8480) and LA-5 (ATCC NO. HB 8479).

Claim 47

The kit of claim 32 wherein said second antibody is the monoclonal antibody produced by the hybridoma Clone C-2 (ATCC No. HB-8478).