Monoclonal antibody analysis of bovine herpesvirus-1 glycoprotein antigenic areas relevant to natural infection

Neutralizing antigenic areas on the glycoproteins of bovine herpesvirus-1 (BHV-1) were identified by reciprocal competition radioimmunoassays using monoclonal antibodies. Three interrelated and two independent antigenic areas were identified on the 77-kDa (K) gIV envelope glycoprotein. Antigenic analysis of this protein has not been previously described. Four interrelated and one independent antigenic areas were found on the 97K gIII envelope glycoprotein. A third group of monoclonal antibodies reacting in Western blot with the 74K subunit of gI, a 130K disulfide-linked 74K/55K heterodimer, revealed four interrelated antigenic areas. All of the antigenic areas on all three glycoproteins were reactive with neutralizing monoclonal antibodies and all were targets for antibody-complement lysis. However, antibodies against gIV were the most efficient at neutralizing the virus and rendering infected cells susceptible to antibody-complement lysis. Convalescent sera from experimentally infected calves were used in a competitive radioimmunoassay to confirm that each antigenic area on the gI, gIII, or gIV glycoproteins was a target for bovine antibodies during primary infection with BHV-1.     

Monoclonal antibodies to bovine coronavirus: characteristics and topographical mapping of neutralizing epitopes on the E2 and E3 glycoproteins

Monoclonal antibodies to the Quebec isolate of bovine coronavirus were produced and characterized. Monoclonal antibodies to both the E2 and the E3 glycoproteins were found to efficiently neutralize virus in vitro. None of the monoclonal antibodies directed against the E1 glycoprotein neutralized virus infectivity. Neutralizing monoclonal antibodies to the E2 glycoprotein were all found to immunoprecipitate gp190, gp100, and their intracellular precursor protein gp170. Neutralizing monoclonal antibodies to the E3 glycoprotein immunoprecipitated gp124 and showed differential reactivity to its precursor proteins gp59 and gp118. These monoclonal antibodies also showed differential reactivity to an apparent degradation product of E3. Neutralizing monoclonal antibodies to E2 bound to two distinct nonoverlapping antigenic domains as defined by competitive binding assays. Neutralizing monoclonal antibodies to the E3 glycoprotein also bound to two distinct antigenic sites as defined by competitive binding assays plus a third site which overlapped these regions. Other results indicated that one domain on the E3 glycoprotein could be further subdivided into two epitopes. Thus four epitopes could be defined by E3-specific monoclonal antibodies.     

Conformation-dependent accessibility of the linear epitopes located on the rabies virus glycoprotein.

Seven monoclonal antibodies (MAbs) were derived from mice immunized with the rabies virus glycoprotein of the Pitman-Moore (PM) strain. These antibodies recognized at least five partially overlapping sites located in one immunodominant region. A panel of MAbs was then used to characterize antigenic relationship between PM strain and SAD-Vnukovo strain of these rabies viruses. In immunoblot, all tested antibodies bound to the glycoprotein of both rabies strains, indicating shared antigenic determinants located on the corresponding immunodominant regions. The pattern of reactivity in immunoblot suggested the specificity of antibodies against linear epitopes. However, the supposed close antigenic relation between PM and SAD-Vnukovo strains (evidenced by immunoblot) was not fully confirmed by immunoenzymatic assay. Data provided by ELISA demonstrated two distinct patterns of MAbs reactivity with both antigens. Four antibodies showed specificity for PM strain glycoprotein only, while three MAbs bound with both PM and SAD-Vnukovo strain antigens. We supposed the strain-specific conformation of the native glycoprotein to be responsible for selective access of single MAbs to the respective common linear epitopes.     

Preparation and characterization of monoclonal antibodies to nucleocapsid protein N and H glycoprotein of rinderpest virus

Fifteen stable mouse spleen cell myeloma hybrids (hybridomas) producing monoclonal antibodies to rinderpest virus proteins were produced. The specificity of these monoclonal antibodies was established by radioimmunoprecipitation followed by polyacrylamide gel analysis and immunofluorescence. Nine antibodies were specific for the surface glycoprotein H. All the nine clones showed inhibition of haemagglutination by measles virus. The antibodies from two clones (A7D2 and B2F6) neutralise infectious virus. Six clones produce antibodies reacting with the nucleocapsid protein N. Three antigenic sites designated I-III, with sites I and II partially overlapping, were topographically mapped on the H molecule by competitive binding assay. Similarly, two antigenic sites I and II were delineated on the N protein. The monoclonal antibodies were used to study the antigenic relationships of H and N proteins of rinderpest virus, measles virus and canine distemper virus.     

Antibodies to distal carboxyl terminal epitopes in the v-fms-coded glycoprotein do not cross-react with the c-fms gene product

The product of the v-fms oncogene is an integral transmembrane glycoprotein that is closely related to the cell surface receptor for the macrophage colony stimulating factor, CSF-1. A fragment of the v-fms gene encoding a major portion of the extracellular amino terminal domain, the membrane-spanning segment, and the entire carboxyl terminal tyrosine kinase domain of the glycoprotein was molecularly cloned into an inducible prokaryotic expression plasmid. Polypeptide products consisting only of v-fms-coded amino acids were produced in bacteria and were used to prepare immune reagents that precipitated the v-fms-coded glycoproteins expressed in transformed cells. Whereas rabbit antisera to recombinant polypeptides detected antigenic determinants of the c-fms proto-oncogene product, seven mouse monoclonal antibodies to these same antigens reacted only with v-fms-specific epitopes. Proteolytic mapping experiments and studies with a mutant v-fms-coded glycoprotein lacking the 37 carboxyl terminal amino acids of the wild-type product showed that the monoclonal antibodies were restricted in their reactivity to epitopes at the extreme carboxyl terminus of the glycoprotein. The v-fms and c-fms gene products must differ significantly in this region.     

Analysis of the S spike (peplomer) glycoprotein of bovine coronavirus synthesized in insect cells.

The bovine coronavirus (BCV) spike glycoprotein precursor (S, formerly termed peplomer) and its two subunit polypeptides (S1 and S2) were individually expressed in Spodoptera frugiperda (Sf9) insect cells. Each recombinant baculovirus expressed both glycosylated (S, 170K; S1, 95K; S2, 80K) and unglycosylated (S0, 140K; S10, 75K; and S20, 65K) forms of BCV spike polypeptides in Sf9 cells. The mature 95K S1 polypeptide was secreted whereas the S and S2 polypeptides remained cell-associated. The S precursor was partially cleaved in Sf9 cells, and the resulting S1 was also released into the medium. Neutralizing monoclonal antibodies representing two antigenic domains bound to recombinant S and S1 but not the S2 polypeptides, indicating that two major epitopes for BCV neutralization are located on the S1 subunit.     

Induction of neutralizing antibody against varicella-zoster virus (VZV) by VZV gp3 and cross-reactivity between VZV gp3 and herpes simplex viruses gB

Glycoprotein gp3 (64K) is one of the major proteins specified by varicella-zoster virus (VZV). This glycoprotein was purified on an immunoadsorbent consisting of monoclonal antibody (clone 8) against gp3 linked to protein A-Sepharose. Rabbits were then immunized with the purified antigen to obtain monospecific antisera against gp3. The monospecific antisera and monoclonal antibody immunoprecipitated polypeptides with the same molecular weights of approximately 64,000 (64K), 106K, and 116K from a lysate of labeled cells infected with VZV. The monoclonal antibodies against gp3 did not have neutralizing activity against VZV, but anti-gp3 monospecific sera neutralized VZV infectivity. The antigenic relation of VZV to herpes simplex virus (HSV) was investigated by the immunofluorescent test, immunoprecipitation followed by analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and the neutralizing antibody test with monoclonal antibodies and monospecific antisera. In the indirect immunofluorescent test, the cytoplasm of cells infected with HSV-type 1 or HSV-type 2 was stained with anti-gp3 monospecific antiserum but not with anti-gp3 monoclonal antibodies. This serum also precipitated the polypeptides of HSV-type 1 and HSV-type 2 with molecular weight of approximately 120,000, possibly corresponding to gB of HSV-1 or HSV-2, and this immunoprecipitation was blocked by anti-gB monoclonal antibody. However, anti-gp3 monospecific antisera did not neutralize either HSV-type 1 or HSV-type 2 infectivity. These results suggest that gp3 induces neutralizing antibody against VZV and that it also has a cross-reacting antigenic determinant with gB of HSV.     

Production and characterization of monoclonal antibodies directed against pseudorabies virus

Hybridoma cell lines producing monoclonal antibodies to pseudorabies virus (PRV) were established. The monoclonal antibodies were characterized with respect to their antigenic specifications and biological activities. Two monoclonal antibodies immunoprecipitated the 50 kDa PRV glycoprotein (gp50) and two immunoprecipitated the 82 kDa glycoprotein (gp82). The monoclonal antibodies were used to analyze the biological roles of these two glycoproteins. One monoclonal antibody directed against each glycoprotein did not require complement for in vitro viral neutralization while the other monoclonal antibody directed against the glycoprotein required complement for neutralization. The monoclonal antibodies against gp50 were shown to be directed against different epitopes within the glycoprotein. In contrast, the monoclonal antibodies against gp82 were shown to be directed against the same antigenic site on the glycoprotein. In vivo passive immunity studies in mice showed that monoclonal antibodies directed against either gp50 or gp82 could be protective.     

Analysis of a glycoprotein of human herpesvirus 6 (HHV-6) using monoclonal antibodies

The virus-specific polypeptides of human herpesvirus 6 (HHV-6) were studied. Six hybridoma clones secreting monoclonal antibodies were established. Localized cytoplasmic antigen and surface antigens were found in and on the infected cells, respectively, by using the immunofluorescent test with those antibodies. In the neutralization test, two clones (OHV3 and OHV9) neutralized HHV-6, but the other five monoclonal antibodies did not. When infected cells were labeled with [35S]methionine and the antigens were immunoprecipitated by OHV3 and OHV9 followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis, two polypeptides with molecular weights of 98,000 (98K) and 92K were detected. Furthermore, it was found that these polypeptides were glycosylated, because they were labeled with [14C]mannose. Pulse-chase studies revealed that precursor molecules were cotranslationaly glycosylated to produce 98K glycoprotein and they were replaced by 92K glycoprotein. Endo-beta-N-acetylglucosaminidases H and F reduced those glycoproteins to putative precursor molecules of 75 kDa. This indicates that N-linked high-mannose sugars associate to 98K and 92K glycoproteins.     

Topographical analysis of epitope relationships on the envelope glycoprotein of yellow fever 17D vaccine and the wild type asibi parent virus

Monoclonal antibodies (MCA), with defined molecular specificity, were used in a competition binding enzyme-linked immunosorbent assay (ELISA) to locate the relative positions of the epitopes on the envelope glycoprotein of yellow fever 17D vaccine virus and its wild type parent virus, Asibi (AS). Five topographically distinct antigenic domains were defined on the E glycoprotein of the 17D vaccine. Three of these (A, B, and C) were represented by one MCA each, a fourth (D) was represented by two MCA, and a fifth domain (E) comprised a major cluster of at least five overlapping epitopes. Asibi virus also possessed domain E which is proposed to be a conserved antigenic region within the envelope glycoprotein of all flaviviruses. Domains A and C were not represented on Asibi virus and one epitope, situated proximal to the E domain, showed structural alterations in physical overlap. Functional activities were assigned to physically mapped epitopes by haemagglutination inhibition (HAI), virus neutralisation (N), and passive protection in mice. The HAI and N functions were not necessarily linked but only MCA with N activity were able to protect mice passively against lethal infection. All domains demonstrated a heterogeneous range of biological properties dependent upon the virus strain rather than the epitope.     

Epitopes functional in neutralization of varicella-zoster virus.

By competition neutralization assay using monoclonal antibodies (MAbs) to varicella-zoster virus (VZV) glycoproteins (gps), we attempted to determine the topographical relationship of epitopes which are functional in VZV neutralization. MAbs against gpI interfered moderately to strongly with neutralization of MAbs against gpIII, and one antigenic domain with two distinct epitopes was identified on gpIII. Competition neutralization assays performed with MAbs to gpI revealed at least three distinct antigenic domains: the first contained two complement-dependent neutralizing epitopes; the second contained five complement-dependent neutralizing, overlapping epitopes and one nonneutralizing, nonoverlapping epitope; and the third contained one complement-enhanced neutralizing epitope. Competition neutralization assays performed with MAbs to gpIV showed one antigenic domain with two distinct epitopes which competed with nonneutralizing gpI MAbs. gpII did not interfere with neutralization of gpI, gpIII, or gpIV. Our data suggest that neutralizing and nonneutralizing MAbs can interfere with the action of viral neutralization either by inhibition or by enhancement. This report describes the epitope mapping of VZV gps by a functional biological assay.     

Characterization of monoclonal antibodies to the hemagglutinin-esterase glycoprotein of a bovine coronavirus associated with winter dysentery and cross-reactivity to field isolates.

Seven hybridoma cell lines producing monoclonal antibodies (MAbs) to the hemagglutinin-esterase (HE) glycoprotein of bovine coronavirus (BCV) were obtained from BALB/c mice that were immunized with an enriched peplomeric fraction of the winter dysentery (WD)-associated strain BCQ.2590. The specificities of these MAbs to either the dimeric (140-kDa) or the monomeric (65-kDa) form of the HE glycoprotein were determined by Western immunoblotting experiments with purified virus and immunoprecipitation tests with [35S]methionine-labelled infected cell extracts. Four of these anti-HE MAbs inhibited the hemagglutinating activity of the virus and three weakly neutralized its infectivity in vitro. In addition, competition binding assays allowed for the definition of two independent antigenic domains (domains A and D) and two overlapping antigenic domains (domains B and C) for the HE glycoprotein of the WD-associated strain; epitopes located within antigenic domain A were not associated with hemagglutination inhibition (HAI) and virus neutralization activities. In HAI tests, the four anti-HA MAbs defined two distinct antigenic subgroups among 24 BCV field isolates that have been associated with either typical outbreaks of WD or neonatal calf diarrhea (NCD) in Quebec dairy herds from 1986 to 1996. The Quebec WD-associated strains of BCV, as well as some of the NCD-associated strains isolated since 1991, fell within a subgroup distinct from that of the prototype Mebus strain.     

Antigenic mapping of the envelope proteins of equine infectious anemia virus: identification of a neutralization domain and a conserved region on glycoprotein 90

Summary Monoclonal antibodies (MCAbs) were used to dissect the antigenic sites of the surface glycoproteins of the prototype cell-adapted Wyoming strain of equine infectious anemia virus (EIAV) Serologic reactivities of these MCAbs were determined by ELISA, additive ELISA, competitive ELISA, and Western blot assays. The results indicated that antigenic reactivity of gp90 was localized on at least four distinct epitopes, two of which were important in neutralization. Our studies also revealed that these epitopes were localized on overlapping antigenic sites on gp90. On the other hand, only two distinct non-overlapping epitopes were identified on gp45. Competitive binding studies of neutralizing MCAbs and reference EIA-positive horse serum delineated the presence of a neutralization domain on gp90 that appears to be immunodominant both in naturally infected horses and in mice immunized with EIAV. Limited proteolytic fragmentation of the gp90 component of several serologically distinct EIAV isolates produced common 12K immunoreactive fragments that contained a conserved epitope. These results indicate the occurrence of conserved antigenic regions on EIAV glycoproteins as well as a neutralization domain on gp90, which can be used as potential targets for vaccine development.     

Characterization of monoclonal antibodies to bovine enteric coronavirus and antigenic variability among Quebec isolates

Summary Twenty monoclonal antibodies (MAbs) were prepared against the Mebus strain of bovine enteric coronavirus, 14 of them reacting with the peplomeric S (gp 100) glycoprotein. Competition binding assays allowed the definition of at least 4 distinct antigenic domains for the S glycoprotein, designated as A, B, C, and D; epitopes associated to neutralizing activity being located in sites A, B, and C. One MAb directed to the hemagglutinin HE (gp 140/gp 65) glycoprotein inhibited the hemagglutinating activity of the virus, but had no neutralizing activity. Comparison of Quebec enteropathogenic BCV isolates using polyclonal antiserum and MAbs directed to the S glycoprotein confirmed their close antigenic relationship, but also revealed the occurrence of at least three distinct antigenic subgroups. Antigenic domain D was highly conserved among BCV isolates, as well as non-neutralizing epitopes assigned to antigenic domains A and C. The Minnesota strain of turkey enteric coronavirus could be distinguished from BCV isolates by MAbs directed to epitopes of antigenic domain C, whereas human coronavirus HCV-OC 43 could be distinguished by MAbs directed to epitopes of antigenic domain A. The porcine hemagglutinating encephalomyelitis virus could be distinguished from the other hemagglutinating coronaviruses by neutralizing epitopes located on antigenic domains A, B, and C.     

Monoclonal antibodies to the HN glycoprotein of newcastle disease virus. Biological characterization and use for strain comparisons

We have isolated nine hybridomas secreting monoclonal antibodies directed against the HN glycoprotein of D₂₆ strain of Newcastle disease virus (NDV) and classified them into three groups by their biological reactivities. The group I antibodies from six clones had high hemagglutination inhibition (HI) and neutralization titers, and exhibited neuraminidase inhibition (NI) activity with substrates, both bovine fetuin and N-acetyl neuraminlactose, although the levels of maximum inhibition were greater with the former substrate. The group II antibodies from two clones also had high HI activity but showed a strongly substrate-dependent NI, with efficient inhibition against fetuin and a total lack of inhibition against neuraminlactose. These antibodies had neutralization capacity but the titers were much lower than those of group I antibodies. One clone (group III secreted antibody which exhibited HI and NI with either substrate but no detectable neutralization activity. Frequency of isolation of antigenic variants and competitive binding assays have shown that group I and group II antibodies may recognize distinct regions not overlapping each other whereas antibodies placed within each of the two groups are to the respective same overlapping regions. The site of group III seemed to be close to or overlap the group I determinant but distinct from group II region. Using the isolated monoclonal antibodies we compared seven heterologous strains by HI and neutralization tests as well as by enzyme-linked immunosorbent assay. The results indicated that the regions recognized by group I antibodies would be conserved among the strains whereas those by the other two antibody groups might undergo considerable changes.     

Monoclonal IgM from patients with peripheral demyelinating neuropathies cross-react with bacterial polypeptides.

Human monoclonal IgM associated with a demyelinating peripheral neuropathy often feature a distinct antibody activity directed against a glucuronyl sulphate epitope shared by myelin-associated glycoprotein (MAG), nerve glycolipids and low molecular weight peripheral nerve polypeptides. Earlier studies showed that these IgM use a diverse repertoire of VH and VL genes which exhibit somatic mutations, possibly indicative of an antigen-driven process. Here, we investigated whether such monoclonal IgM may react with environmental bacterial antigens. We found that six patients' sera and purified monoclonal IgM, as well as IgM from supernatants of three clonal anti-MAG-secreting cell lines reacted with unique 90-100 kD polypeptides from extracts of two out of 10 bacterial species. Purified MAG was able to inhibit this reactivity. These results indicate molecular mimicry as a possible mechanism of this immunomediated neuropathy and associated clonal lymphoid disease.     

Characterization of polypeptides in Rickettsia tsutsugamushi: effect of preparative conditions on migration of polypeptides in polyacrylamide gel electrophoresis.

The polypeptide compositions and antigenic components of Rickettsia tsutsugamushi were analyzed by modifying the solubilization conditions prior to polyacrylamide gel electrophoresis and by using monoclonal antibodies in immunoblotting experiments. Several polypeptides were converted to larger or smaller molecules by using various conditions for rickettsial sample preparation. Solubilization of a sample in 2-mercaptoethanol-containing buffer resulted in conversion of high-molecular-weight polypeptides to smaller polypeptides and conversion of some of the 43-kilodalton (43K) polypeptide to a 46K polypeptide. The heat modifiability of selected polypeptides was shown by heating samples at 100 degrees C. A major polypeptide on the rickettsial surface which showed strain-specific antigenicity appeared at the 43K position in samples solubilized at 37 degrees C but moved to the 56K position after samples were heated at 100 degrees C. Immunoblotting with an anti-56K polypeptide monoclonal antibody demonstrated that the reactive antigens existed predominantly as the higher-molecular-weight polypeptides. These polypeptides were converted to 43K polypeptides at 37 degrees C or the 56K polypeptides at 100 degrees C by cleavage of disulfide linkages with 2-mercaptoethanol treatment.     

Utilization of anion-exchange chromatography and monoclonal antibodies to characterize multiple pilus types on a uropathogenic Escherichia coli O6 isolate.

Multiple pilus types from a uropathogenic strain of Escherichia coli O6, strain 6260, were characterized by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), high-pressure liquid chromatography, binding assays, and erythrocyte adsorption. In addition, monoclonal antibodies were raised against purified pili of E. coli 6260 and used for immunological characterization. SDS-PAGE analysis of the purified pili showed at least three different subunits with molecular weights of 15,700, 17,800, and 19,300. SDS-PAGE analysis of four protein peaks from anion-exchange chromatography of intact pili showed polypeptides with molecular weights of 19,300 (fraction 1), 15,700 (fraction 2), and 17,800 and 15,700 (both fractions 3 and 4). Erythrocyte adsorption of the whole-pilus preparation removed the 17,800-molecular-weight subunit (17.8K subunit) and reduced the 15.7K subunit. Pili from an isogenic hemagglutination-negative variant of E. coli 6260, showing only the 15.7K and 19.3K subunits by SDS-PAGE, lacked the 17.8K subunit of fractions 3 and 4 present in the parent high-pressure liquid chromatography profile. Our data suggest that two of the pilus subunits, the 15.7K and 17.8K subunits, mediate mannose-resistant agglutination of human erythrocytes. Pili in fractions 1 and 2 from the parent strain bound specifically to mannose residues, while pili in fraction 4 bound to P-coated horse erythrocytes; no receptor specificity was identified for pili in fraction 3. Immunological analysis by the immunoblot technique showed that monoclonal antibody 11-2 reacted with the 19.3K subunit, monoclonal antibodies 34-3 and 73-3 reacted with the 15.7K subunit, and monoclonal antibodies 81-1, 82-1, and 91-1 reacted with polymers of subunits retained in the stacking gel. Intact pili precipitated by any of the six monoclonal antibodies showed two polypeptides by SDS-PAGE: 15.7K and 19.3K polypeptides for monoclonal antibody 11-2, and 15.7K and 17.8K polypeptides for monoclonal antibodies 34-3, 73-3, 81-1, 82-1, and 91-1. The cross-reactivity of the monoclonal antibodies with purified pili from other E. coli strains was determined by enzyme-linked immunosorbent assay. Monoclonal antibody 11-2 showed no significant cross-reactivity with heterogeneous pili. In contrast, the other monoclonal antibodies showed equivalent or greater reactivity with P pili from heterologous strains as compared with reactivity with E. coli 6260 pili.(ABSTRACT TRUNCATED AT 400 WORDS)     

Fine mapping of a peptide sequence containing an antigenic site conserved among arenaviruses

The lymphocytic choriomeningitis virus (LCMV) structural glycoproteins GP-1 (Mr 44K) and GP-2 (Mr 35K) are encoded on a single intracellular proteolytic cleavage precursor glycoprotein, GP-C (Mr 76K). We have used a series of synthetic peptides derived from the deduced amino acid sequence of LCMV GP-C to define an antigenic site containing two topographically overlapping epitopes. Three mouse monoclonal antibodies directed against two epitopes on GP-2 were assayed for binding in solution phase blocking and solid-phase enzyme-linked immunoadsorbant assays to a series of peptides representing the sequence of the intracellular precursor glycopeptide GP-C. Both epitopes were initially localized to a single peptide GP-C 370-382 (Cys-Asn-Tyr-Ser-Lys-Phe-Trp-Tyr-Leu-Glu-His-Ala-Lys) in the GP-2 segment of GP-C. Further analysis demonstrated that both epitopes were contained within a nine amino acid segment, GP-C 370-378, which contains five residues conserved among LCMV, Lassa, Pichinde, and Tacaribe viruses. Assays with N-terminal deletions from this sequence suggested that the minimal epitope recognized by the broadly cross-reactive monoclonal 33.6 (epitope GP-2a) consisted of five amino acids, GP-C 374-378 (Lys-Phe-Trp-Tyr-Leu). Reactivity of a second monoclonal, 9-7.9 (epitope GP-2B) but not 33.6, was abolished when substitution of tyrosine for phenylalanine was made at position 375 in the antigenic sequence corresponding to a naturally occurring sequence difference between LCM and Lassa viruses. Polyclonal sera from human cases and from animals experimentally infected with Junin, LCM, and Lassa viruses, respectively, bound to the antigenic peptide GP-C 370-382 but not to control peptides. As was the case with the monoclonals, this binding activity was abrogated by blocking with the antigenic peptide but not with control peptides in solution.     

Monoclonal antibody-mediated immunoprecipitation of proteins from cell,infected with marek's disease virus or turkey herpesvirus

The major immunogenic viral proteins of Marek's disease virus (MDV) and turkey herpesvirus (HVT) share antigenic determinants. The polyacrylamide gel electrophoresis pattern of five viral polypeptides immunoprecipitated with homologous convalescent chicken plasma was identical with the pattern obtained after immunoprecipitation with heterologous convalescent chicken plasma. A panel of monoclonal antibodies was used to identify MDV and HVT polypeptides. Sixteen monoclonal antibodies were positive in an immunofiuorescence assay. However, only eight monoclonal antibodies immunoprecipitated a total of seven distinct viral proteins from MDV- and HVT-infected cells. Six of these monoclonals immunoprecipitated multiple viral proteins. None of the monoclonal antibodies recognized the common A antigen of MDV or HVT. Monoclonal antibodies immunoprecipitated three glycoproteins (100,000, 60,000, and 49,000 Da) that comigrate in polyacrylamide gels with three of the five common polypeptides obtained with the convalescent chicken plasma. In addition, a 79,000-Da protein was common to all MDV- and HVT-infected cells. Competition immunoprecipitation and peptide mapping by limited proteolysis confirmed that the three glycoproteins and the 79,000-Da protein contain MDV-HVT common epitopes. MDV-specific antigenic determinants were detected on the three remaining viral proteins (41,000, 38,000, and 24,000 Da).