A METHOD OF DETERMINING TYPE SPECIFIC ANTIBODIES TO HERPES SIMPLEX VIRUS TYPES 1 AND 2

Abstract: This enclosure concerns a method of serotyping antibodies to HSV 2 or 1 or in a sample suspected to contain HSV antibodies which comprises: a) reacting the sample and at least one monoclonal antibody specific for HSV-2 or HSV-1 glycoprotein G with an antigen wherein said monoclonal antibody is not susceptible to heterologous blocking; and b) determining whether antibody binding has occurred. The serotyping method of the disclosure can be done in a variety of formats such as sequential or simultaneous. In another embodiment, this disclosure concerns kits for serotyping antibodies to HSV-1 and HSV-2.

Description: HI£ A METHOD OF DETERMINING TYPE SPECIFIC ANTIBODIES TO HERPES SIMPLEX VIRUS TΪPES 1 AND 2 FIELD OF THE INVENTION This invention concerns a method of serotyping antibodies to herpes simplex virus types 1 and 2, and, in particular, to a method of serotyping which employs monoclonal antibodies to HSV-1 gG and HSV-2 gG which are not susceptible to heterologous blocking. BACKGROUND OF THE INVENTION The most striking characteristic of herpes simplex virus (HSV) is its propensity for persisting in a quiescent or latent state in man, with recurrence of activity at irregular intervals. There are two main immunologic variants, types 1 and 2, which crossreact. One of the major problems in seroepidemiologic studies of HSV infection is the differentiation of antibodies in human serum directed against HSV-1 and HSV-2. Researchers have attempted to overcome these problems using purified HSV glycoproteins and absorption to remove cross reacting antibodies in an effort to develop a rapid method of distinguishing HSV-1 and HSV-2 antibodies. Such methods have important clinical and epide iological significance because it permits diagnosis of persons with asymptomatic HSV infection, identification of candidates for antiviral therapy, as well as identification of pregnant women who may transmit a potentially fatal HSV infection to the newborn. U.S. Patent 4,764,459, issued to Hampar et al. on August 16, 1988, and Hampar et al., J. Clin. Microbiol., 21: 496-500 (1985), describe an ELISA to determine antibodies against HSV types 1 and 2 in human sera. The method utilizes heterologous virus-infected cell extracts to absorb human sera to remove intertypic cross-reactive antibodies. Specifically, the sera were absorbed with heterologous virus-infected cell extracts- to remove cross-reacting antibodies and then were applied to ELISA plates containing the target antigens, immunoaffinity-purified HSV-1 glycoproteins gC and gD and HSV-2 glycoproteins gD and gF. The absorbance index, defined as the ratio of the absorbance generated by a serum sample absorbed with a heterologous virus- infected cell extract versus the absorbance generated by a serum sample absorbed with an uninfected cell extract, was used to determine the presence or absence of antibodies to HSV-1 and HSV-2. U.S. Patent 4,855,224, issued to Berman et al. on August 8, 1989, describes the use of a molecularly cloned polypeptide with antigenic determinants capable of specifically binding complementary antibody. Using an HSV-1 glycoprotein D as the polypeptide, HSV antibodies could be determined. HSV-2 antibodies could be detected using HSV-2 glycoprotein C fragment. Arvin et al.. Infection and Immunity, 40: 184-189 (1983) , describe an immunoaffinity purification of HSV-1 glycoprotein C with a monoclonal antibody and the use of this purified glycoprotein in a solid phase RIA for type-specific antibodies to HSV-1. Svennerhol et al., J. Clin. Microbiol., 19: 235-239 (February 1984), describe the use of helix pomatia lectin purified HSV-1 and HSV-2 antigens to determine type specific antibodies in human sera. Identification of the lialix nma ia lectin purified HSV antigens (HSV-1 gC and HSV-2 gG) was described by Olofsson et al., J. gen. Virol., 67: 737-744 (1986). Lee et al., J. Clin. Microbiol., 22: 641-644 (1985), describe the evaluation of an HSV-2 type- specific enzyme-linked immunodot serological assay based on affinity-purified gG-2 as antigen. Lee et al., J. Virol. Methods, 14: 111-118 (1986), describe immunoaffinity purification of HSV-1 glycoprotein G using a monoclonal antibody. The purified antigen was coated on a solid phase in an immunodot enzymatic assay to detect HSV-1 antibodies. Ross et al., J. Virology, 54: 851-855 (1985), describe a competitive ELISA to test for human antibodies to antigenic sites on HSV glycoproteins C and D, which are recognized by mouse monoclonal antibodies. It was shown that there were antibodies in human sera that were capable of blocking the binding of monoclonal antibodies. Thus, even though the monoclonal antibodies used were specific for HSV-1 or HSV-2 the assay could not be used to distinguish HSV-1 and HSV-2 antibodies because blocking by heterologous human antibodies occurred, i.e., human antibodies to HSV-1 blocked monoclonal antibodies specific for HSV-2 and human antibodies to HSV-2 blocked monoclonal antibodies specific for HSV-1. SUMMARY OF THE INVENTION This invention concerns a method of serotyping antibodies to HSV 2 or 1 or in a sample suspected to contain HSV antibodies which comprises: a) reacting the sample and at least one monoclonal antibody specific for HSV-2 or HSV-1 glycoprotein G with an antigen wherein said monoclonal antibody is not susceptible to heterologous blocking; and b) determining whether antibody binding has occurred. The serotyping method of the invention can be done in a variety of formats such as sequential or simultaneous. In another embodiment, this invention concerns kits for serotyping antibodies to HSV-1 and HSV-2. BRIEF DESCRIPTION OF THE DRAWINGS Figure 1 is a graph depicting the absorbance values for an assay used to determine anti-HSV-1 antibodies. Figure 2 is a graph depicting the absorbance values for an assay to determine HSV-2 antibodies. DETAILED DESCRIPTION OF THE INVENTION The term "antigen" refers to an antigenic substance which can be natural, recombinant, or synthetic, and which is capable of binding to the monoclonal antibodies and to the antibodies in the sample. The antigen need not be substantially pure as long as the contaminants do not interfere with the assay method. The term HSV-2 glycoprotein G (HSV-2 gG) refers to the HSV-2 glycoprotein as described by Roizman et al., Virology 133 242-247, 1984; Marsden et al., J. Virology 50. 547-554, 1984; Lee et al., J. Clin. Microbiol. 22., 641-644, 1985; Balachandran et al., J. Virology $£, 825-832, 1985; and Su et al., J. Virology £2., 3668-3674, 1988. The term HSV-1 glycoprotein G (HSV-1 gG) refers to the HSV-1 glycoprotein as described by Ackermann et al., Virology 150. 207-220, 1986; Richman et al., J. Virology 5L, 647-655, 1986; and Sullivan et al., J. Gen. Virol. £&, 2587-2598, 1987. The assay of the present invention is an extraordinarily simple and specific method to serotype antibodies to HSV-2 or HSV-1 which overcomes the cross- reactivity problems which have plagued researchers in differentiating these antibodies. HSV-1 antibodies are serotyped by using at least one monoclonal antibody specific for HSV-1 glycoprotein G which is not susceptible to heterologous blocking. HSV-2 antibodies are serotyped by using at least one monoclonal antibody specific for HSV-2 glycoprotein G which is not susceptible to heterologous blocking. These monoclonal antibodies can be unlabeled or labeled as is discussed below. The following is one illustration of the method of the invention: HSV antigen, types 1 and 2, is immobilized on a suitable support. It is then reacted with a sample, usually serum or plasma, which is suspected to contain HSV antibodies which, if present, will bind to the antigen, and, thus, block binding of the monoclonal antibody. After excess reagents are removed by washing, a labeled monoclonal antibody is incubated with the mixture. Antibodies to HSV-1 are detected by using a monoclonal antibody specific for HSV-1 glycoprotein G . Antibodies to HSV-2 are detected by using a monoclonal antibody specific for HSV-2 glycoprotein G. The mixture is washed again and antibody binding is determined by measuring the amount of reporter bound to the solid support. The sample need not be limited to serum or plasma. Any biological fluid suspected to contain HSV antibodies can be used as the sample. Techniques for preparing monoclonal antibodies which can be used to practice the invention are well known. The preparation of monoclonal antibodies which recognize either HSV-1 or HSV-2 glycoprotein G have been described in a variety of publications such as Roizman et al.. Virology 133. 242-247, 1984; Marsden et al., J. Virology 5_£, 547-554, 1984-; Balachandran et al., J. Virology 5_A, 825-832, 1985; Lee et al., J. Clin. Microbiol. 22., 641-644, 1985; Ackermann et al.. Virology 150 P 207-220, 1986; and Richman et al., J. Virology 51, 647-655, 1986. The antibody can be labeled with a reporter or with a member of an immune or non-immune specific binding pair. When the latter is used, detection is effected by reacting the complex with the other member of the binding pair which is labeled. Members of specific binding pairs suitable for practicing the invention can be of the immune or non- immune type. Examples of immune specific binding pairs include antigen-antibody systems or hapten-anti-hapten systems. The antibody member of the specific binding pair can be produced by customary methods familiar to those skilled in the art. Such methods involve immunizing an animal with the antigen member of the specific binding pair. If the antigen member of the specific binding pair is not immunogenic, i.e., a hapten, it can be covalently coupled to a carrier protein to render it immunogenic. The antibody member whether polyclonal, monoclonal, or an immunoreactive fragment thereof can be produced by conventional techniques well known to those skilled in the art. The terms immunoreactive antibody fragment or immunoreactive fragment mean fragments which contain the binding region of the antibody. Such fragments can be Fab-type fragments which are defined as fragments devoid of the Fc portion, e.g.. Fab, Fab' and F(ab') 2 fragments, or may be so-called "half-molecule" fragments obtained by reductive cleavage of the disulfide bonds connecting the heavy chain components of the intact antibody. Non-immune binding pairs include systems wherein the two components share a natural affinity for each other but are not antibodies. Exemplary non-immune binding pairs are biotin-avidin or biotin-streptavidin, folic acid-folate binding protein, complementary probe nucleic acids, etc. A variety of methods are available to covalently label monoclonal antibodies with members of specific binding pairs. Methods are selected based upon the nature of the member of the specific binding, pair, the type of linkage desired, and the tolerance of the antibody to various conjugation chemistries. Biotin can be covalently coupled to monoclonal antibodies by utilizing commercially available active derivatives. Some of these are biotin-N-hydroxysuccinimide which binds to amine groups on proteins; biotin hydrazide which binds to carbohydrate moieties, aldehydes and carboxyl groups via a carbodiimide coupling; and biotin maleimide and iodoacetyl biotin which bind to sulfhydryl groups. Fluorescein can be coupled to protein amine groups using fluorescein isothiocyanate. Dinitrophenyl groups can be coupled to protein amine groups using 2,4-dinitrobenzene sulfonate or 2,4-dinitrofluoro- benzene. Other standard methods of conjugate may be employed to couple monoclonal antibodies to a member of a specific binding pair including dialdehyde, carbodiimide coupling, homofunctional crosslinking, and heterobifunctional crosslinking. Carbodiimide coupling is an effective method of coupling carboxyl groups on one substance to amine groups on another. Carbodiimide coupling is facilitated by using the commercially available reagent, l-ethyl-3-(dimethylaminopropyl)- carbodiimide (EDAC) . Homobifunctional crosslinkers, including the bifunctional i idoesters and bifunctional N-hydroxy- succinimide esters, are commercially available and are employed for coupling amine groups on one substance to amine groups on another. Heterobifunctional crosslinkers are reagents which possess different functional groups. The most common commercially available heterobifunctional crosslinkers have an amine reactive N-hydroxysuccinimide ester as one functional group, and a sulfhydryl reactive group as the second functional group. The most common sulfhydryl reactive groups are maleimides, pyridyl disulfides and active halogens. One of the functional groups may be a photoactive aryl nitrene, which upon irradiation reacts with a variety of groups. A reporter can be linked directly or indirectly, covalently or non-covalently to the monoclonal antibodies or to a member of a specific binding pair. Reporters can be radioactive isotopes, enzymes, fluorogenic, chemiluminescent or electrochemical materials. Two commonly used radioisotopes are 125 I and 3 H. Standard radioactive isotopic labeling procedures include the chloramine T, lactoperoxidase and Bolton- Hunter methods for 125 I and reductive methylation for 3 H. Enzymes are also used as reporters for immunoassays. These include, but are not limited to, horseradish peroxidase, alkaline phosphatase, β-galactosidase, glucose oxidase, luciferase, β-lactamase, urease and lysozyme. Labeling with enzymes is facilitated by using dialdehyde, carbodiimide coupling, homobifunctional crosslinkers and heterobifunctional crosslinkers as described above for labeling monoclonal antibodies with members of specific binding pairs. The labeling method chosen depends on the functional groups available on the enzyme and the material to be labeled, and the tolerance of both to the conjugation conditions. The labeling method used in the present invention may be one of, but not limited to, any conventional methods currently employed including those described by Engvall and Pearlmann, Immunochemistry 8, 871 (1971), Avrameas and Ternynck, Immunochemistry 8, 1175 (1971), Ishikawa et al., J. Immunoassay 4(3):209- 327 (1983) and Jablonski, Anal. Biochem. 148:199 (1985). Labeling may be accomplished by indirect methods such as using spacers or other members of specific binding pairs. .An example of this is the detection of biotinylated antibodies with unlabeled streptavidin and biotinylated enzyme, with the unlabeled streptavidin and biotinylated enzyme being added either sequentially or simultaneously. Detection of enzyme activity may be facilitated by measuring chromogenic, fluorogenic, che iluminescent or electrochemical changes by commonly known methods. Reporters can be fluorogenic or chemiluminescent in nature. In addition, reporters can be detectable by electrochemical means. Some methods of labeling with these reporters are described above. In a preferred embodiment, the enzyme, horseradish peroxidase is used as the reporter. In another variation, the monoclonal antibody need not be labeled. If it is unlabeled, it can be detected using a labeled anti-species immunoglobulin reagent. If more than one monoclonal antibody is used, the combination should be specific for either HSV-1 gG or HSV-2 gG. Another embodiment involves the simultaneous, rather than sequential, addition of sample and monoclonal antibody to a support containing immobilized antigen. This is illustrated in the examples described below. Alternatively, at least one monoclonal antibody can be immobilized on a support. It can then be reacted with labeled antigen and sample or the labeled antigen and sample can be preincubated and then reacted with the support containing immobilized monoclonal antibody. Suitable supports include synthetic polymer supports, such as polystyrene, polypropylene, substituted polystyrene, e.g., aminated or carboxylated polystyrene; polyacrylamides; polyamides; polyvinyl- chloride, etc.; glass beads; agarose; nitrocellulose, etc. 10 This invention also concerns a kit for serotyping antibodies to HSV-1 or HSV-2 or both HSV-1 and HSV-2. A kit according to the present invention comprises (a) a support containing immobilized HSV antigen, (b) a quantity of at least one monoclonal antibody specific for HSV-1, HSV-2 or a combination thereof wherein said antibody is not susceptible to heterologous blocking and further wherein said antibody is either (i) unlabeled, (ii) labeled with a reporter, or (iii) labeled with a first member of a specific binding pair, and (c) a quantity of labeled anti-antibody when the monoclonal antibody is unlabeled or a quantity of a labeled second member of the binding pair when the monoclonal antibody is labeled with the first member of the specific binding pair. The following examples are intended to illustrate the invention. EXAMPLE 1 Blocking of Anti-HSV 2 gG Monoclonal Antibodies by Serum Containing ftnti HSV-2 Antibodies Preparation of Monoclonal Antibodies The monoclonal antibodies were prepared according to the methods of L. Pereira et al., as described in Infection and Immunity, Vol. 29, No. 2, pp. 724-732 (Aug. 1980) and Oi, et al., as described in Immunoglobulin Producing Hybrid Cell Lines, pp. 351-371, in B. Mishell and S. Schiigi (ed.). Selected Methods in Cellular Immunology, W. H. Freeman Co., San Francisco. Monoclonal antibodies were purified from ascites fluid by salt fractionation with 40% saturated ammonium sulfate and gel filtration on Sephacryl S-300 (Pharmacia, Inc., Piscataway, NJ) . Purified monoclonal antibodies were labeled with horseradish peroxidase (HRP) according to the SMCC method described by Ishikawa 11 et al.. Journal of Immunoassay, Vol. 4, No. 3, pp. 209-327, 1983. Polystyrene microplate wells were coated with HSV-2 antigen (Scripps Laboratories, San Diego, CA) diluted in 0.1M carbonate buffer pH 9.6, and blocked with 2% bovine serum albumin (BSA) in 10 mM phosphate buffered saline (PBS), pH 7.4. For the sequential (Seq.) format, a serum specimen containing antibodies to HSV-2 (Pos.) or a specimen without antibodies to HSV (Neg.) were incubated for 1 hr at 37°C. Following washing with PBS containing 0.05% Tween 20 (PBST) , HRP labeled anti-HSV-2 gG monoclonal antibodies 55311, 55312, or 55311 and 55312 combined at 1:1, diluted in 1% BSA-PBST were incubated for 1 hr at 37°C. After washing with PBST, a tetramethylbenzidine (TMB) substrate (SOMA Laboratories, Romeo, MI) was added for 30 min., the reaction stopped with H 2 SO 4 , and the absorbance at 450 nm read on a microplate reader (Vmax, Molecular Devices Corp., Menlo Park, CA) . For the simultaneous (Sim.) format, the monoclonal antibodies were diluted into the serum specimens, added to the wells, and incubated for 1 hr at 37°C. Following washing with PBST, TMB was added and absorbance determined as described for the sequential format. Results The results in Table I below indicate that the anti-HSV-2 containing serum blocked the binding of both anti-HSV-2 gG monoclonal a'ntibodies when used alone, or in combination, whether in a sequential or simultaneous assay format. 12 Table I Blocking of Anti-HSV 2 gG Monoclonal Antibodies by Serum Containing Anti-HSV 2 Antibodies Antibody O.D. 55311 1.441 0.193 55312 0.999 0.010 55311 + 55312 1.510 0.098 55311 0.836 0.000 55312 1.033 0.000 55311 + 55312 1.195 0.000 Polystyrene microplate wells were coated with a combination of HSV-1 and HSV-2 antigens (Scripps Laboratories, San Diego, CA) as described in Example 1 above. For each of the two specificity determinations, twenty serum samples containing antibodies to HSV-1 and twenty serum samples containing antibodies to HSV-2, diluted 1:1 with 1% BSA-PBS-0.5% triton X-100, were added to the wells, incubated for 1 hr at 37°C, and washed with PBST. To determine specificity to anti-HSV-2, HRP labeled monoclonal antibody 55312 (anti-HSV-2 gG) diluted in 1% BSA-PBST, was added for 1 hr at 37°C. To determine specificity to anti-HSV-1, HRP labeled monoclonal antibody 55315 (anti-HSV-1 gG) diluted in 1% BSA-PBST, was added for 1 hr at 37°C. Following washing with PBST, TMB was added, and the absorbance determined as described in Example 1. 13 Results Figure 1 is a graph showing the absorbance values for the twenty anti-HSV-1 and twenty anti-HSV-2 antibody containing sera used in an assay to determine anti-HSV-1 antibodies. The sera containing anti-HSV-1 antibodies blocked the binding of the anti-HSV-1 gG monoclonal antibody. No such blocking was observed with the sera containing anti-HSV-2 antibodies. Figure 2 is a graph showing the absorbance values obtained for an assay to determine anti-HSV-2 antibodies. The sera containing anti-HSV-2 antibodies blocked the binding of the anti-HSV-2 gG monoclonal antibody. No such blocking was observed with the sera containing anti-HSV-1 antibodies. EXAMPLE 3 Typing Anti-HSV Antibodies in Serum Specimens Polystyrene microplate wells were coated with a combination of HSV-1 and HSV-2 antigens as described in Example 1 above. Twenty serum samples containing antibodies to HSV and one negative serum sample were added to the wells in duplicate for 1 hr at 37°C, and washed with PBST. To one of the duplicate wells, HRP labeled 55312 (anti-HSV-2 gG) was added, and to the other, HRP labeled 55315 (anti-HSV-1 gG) was added. Labeled antibodies were diluted in 1% BSA-PBST, and incubated for 1 hr at 37°C. Following washing with PBST, TMB was added, and the absorbance determined as described in Example 1. Results The absorbances are shown in Table II. A positive response was chosen as at least a 50% reduction in the absorbance of the negative control specimen. Determining anti-HSV types by this method does not require tedious antigen purification procedures, heterologous absorption methods, or ratio determinations as all the previously described methods require. TABLE II Typing Anti-HSV Antibodies in Serum Specimens Absorbance 450 nm

Claims: 15 What is claimed is: 1. A method of typing antibodies to HSV 2 in a sample suspected to contain HSV antibodies which comprises: a) reacting the sample and at least one monoclonal antibody specific for HSV-2 glycoprotein G with an antigen wherein said monoclonal antibody is not susceptible to heterologous blocking; and b) determining whether antibody binding has occurred. 2. A method of typing antibodies to HSV 1 in a sample suspected to contain HSV antibodies which comprises: a) reacting the sample and at least one monoclonal antibody specific for HSV-1 glycoprotein G with an antigen wherein said monoclonal antibody is not susceptible to heterologous blocking; and b) determining whether antibody binding has occurred. 3. A method of typing antibodies to HSV-2 in a sample suspected to contain HSV antibodies which comprises: a) contacting a support containing immobilized antigen with the sample; b) reacting the product of step (a) with at least one monoclonal antibody specific for HSV-2 glycoprotein G wherein said antibody is not susceptible to heterologous blocking and further wherein said antibody is either (i) unlabeled, (ii) labeled with a reporter, or (iii) labeled with a first member of a specific binding pair; 16 c) reacting the product of step (b) (i) with a labeled anti-antibody or reacting the product of step (b) (iii) with a labeled second member of the specific binding pair; and d) detecting the presence or absence of label. 4. A method according to claim 3 wherein the antibody is labeled with horseradish peroxidase. 5. A method according to claim 4 wherein the horseradish peroxidase labeled antibody is reacted with tetra eth lbenzidene. 6. A method of serotyping antibodies to HSV-1 in a sample suspected to contain HSV antibodies which comprises: a) contacting a support containing immobilized antigen with the sample; b) reacting the product of step (a) with at least one monoclonal antibody specific for HSV-1 glycoprotein G wherein said antibody is not susceptible to heterologous blocking and further wherein said antibody is either (i) unlabeled, (ii) labeled with a reporter, or <iii) labeled with a first member of a specific binding pair; c) reacting the product of step (b) (i) with a labeled anti-antibody or reacting the product of step (b) (iii) with a labeled second member of the specific binding pair; and d) detecting the presence or absence of label. 7. A method according to claim 6 wherein the antibody is labeled with horseradish peroxidase. 17 8. A method according to claim 7 wherein the horseradish peroxidase labeled antibody is reacted with tetramethylbenzidene. 9. A method according to claim 3 wherein steps (a) and (b) are performed simultaneously. 10. A method according to claim 6 wherein steps (a) and (b) are performed simultaneously. 11. A method of typing antibodies to HSV-2 in a sample suspected to contain HSV antibodies which comprises: a) contacting a support containing at least one immobilized monoclonal antibody specific for HSV-2 glycoprotein G with labeled antigen and the sample wherein said monoclonal antibody is not susceptible to heterologous blocking; and b) detecting the presence or absence of label. 12. A method of typing antibodies to HSV-1 in a sample suspected to contain HSV antibodies which comprises: a) contacting a support containing at least one immobilized monoclonal antibody specific for HSV-1 glycoprotein G with labeled antigen and the sample wherein said monoclonal antibody is not susceptible to heterologous blocking; and b) detecting the presence or absence of label. 18 13. A method of typing antibodies to HSV-2 in a sample suspected to contain HSV antibodies which comprises: a) preincubating the sample with labeled antigen; b) reacting the product of step (a) with a support containing at least one immobilized monoclonal antibody specific for HSV-2 glycoprotein G wherein said antibody is not susceptible to heterologous blocking; and c) detecting the presence or absence of label. 14. A method of serotyping antibodies to HSV-1 in a sample suspected to contain HSV antibodies which comprises: a) preincubating the sample with labeled antigen; b) reacting the product of step (a) with a support containing at least one immobilized monoclonal antibody specific for HSV-1 glycoprotein G wherein said antibody is not susceptible to heterologous blocking; and c) detecting the presence or absence of label. 15. A kit of reagents for serotyping antibodies to HSV-1, HSV-2, or a combination thereof which comprises: a) a support containing immobilized HSV antigen; b) a quantity of at least one monoclonal antibody specific for HSV-1, HSV-2 or a combination thereof wherein said antibody is not susceptible to heterologous blocking and further wherein said antibody is either (i) unlabeled, (ii) labeled with a reporter. 19 or (iii) labeled with a first member of a specific binding pair; and c) a quantity of labeled anti-antibody when the monoclonal antibody is unlabeled or a quantity of a labeled second member of a binding pair when the monoclonal antibody is labeled with the first member of a specific binding pair. 16. A kit according to claim 15 wherein the antibody is labeled with horseradish peroxidase.

Inventors: BOBROW MARK NORMAN, BOBROW, MARK, NORMAN
Applicants: DU PONT, E.I. DU PONT DE NEMOURS AND COMPANY
Application year: 1991
Main classification: G01N33
IPC codes: G01N33