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The present study envisages expression of immunodominant ectodomain of peste des petits ruminants virus (PPRV) fusion (F) protein in Escherichia coli BL21 (DE3) and its characterization to assess its immunoreactivity. The ectodomain gene sequences corresponding to 222 amino acids, was amplified from PPR vaccine virus, cloned into pET33b vector and expressed in E. coli at an optimal temperature of 37 degrees C with 1 mM IPTG for 5 h. The expressed and Ni-NTA purified PPRV F protein (31 kDa) was characterized by SDS-PAGE and Western blot using anti-his-tagged-conjugate, anti-serum raised against recombinant PPRV F protein, hyper immune serum against whole PPRV and convalescent sera from sheep and goats. The expressed protein was assessed for its immunoreactivity by ELISA and immunoblotting. The antibody response mounted against the recombinant PPRV F protein in immunized rabbits was detected by recombinant PPRV F antigen based indirect ELISA, and whole virus antigen based indirect ELISA, which indicating the native confirmation of the expressed protein in E. coli. Indirect ELISA was optimized using known true positive and negative sera with respect to PPRV antibodies in order to assess the reactivity of the PPRV F protein in detecting PPRV F antibodies in small ruminants. The E. coli expressed recombinant ectodomain of PPRV F protein exhibits immunoreactivity and was able to specifically detect PPRV antibodies in response to both vaccination and disease in natural host. (C) 2016 Elsevier B.V. All rights reserved.

Post-transcriptional modification of viral mRNA is essential for the translation of viral proteins by cellular translation machinery. Due to the cytoplasmic replication of Paramyxoviruses, the viral-encoded RNA-dependent RNA polymerase (RdRP) is thought to possess all activities required for mRNA capping and methylation. In the present work, using partially purified recombinant RNA polymerase complex of rinderpest virus expressed in insect cells, we demonstrate the in vitro methylation of capped mRNA. Further, we show that a recombinant C-terminal fragment (1717-2183 aa) of L protein is capable of methylating capped mRNA, suggesting that the various post-transcriptional activities of the L protein are located in independently folding domains.

Peste des Petits Ruminants (PPR) is a highly contagious transboundary animal disease of wild and domestic small ruminants and notifiable byworld organization for animal health (OIE). It constitutes a serious threat to livestock production in many developing countries, in western Africa and southern Asia. The disease is caused by peste des petits ruminants virus (PPRV), a negative sense, single stranded RNAvirus belonging to the genus Morbillivirus of the family Paramyxoviridae (Gibbs et al. 1979). PPRV causes huge disease burden on agriculture across developing world, affecting small ruminant production and in turn increases poverty in some of the poorest parts of the world (Banyard et al. 2010). In PPR endemic regions of the world, conventional live attenuated vaccines are being used to vaccinate small ruminants (Diallo et al. 1989). Currently, the original Nigerian strain of PPRV (Nig 75/1) is used in the African countries (Diallo 2003). In India, three Indian strains of PPRV have been attenuated and shown to provide protective immunity (Saravanan et al. 2010). Although the live attenuated vaccines are efficacious, they suffer from the same draw back as that of live attenuated Rinderpest vaccine in terms of heat stability. This puts severe constraints on the coverage of small ruminant’s population during vaccination campaigns. Although there is one report of a method for making a heat stable live attenuated vaccine (Worrwall et al. 2000), such a thermo stable vaccine has not been put to use or field tested. Another major disadvantage of using live attenuated vaccine is that the antibody responses they induce in animals cannot be distinguished from those following a natural infection. This makes sero-epidemio surveillance of the disease impossible in endemic areas where a vaccination programme has been or is being implemented. The immune response to a given antigen can be regulated by a number of idiotypic determinants (Ids), and its counterpart, an anti-idiotypic antibody (anti-Id) or Ab2 (Jerne 1974). An idiotype is located on the variable regions of antibody molecules. An internal image anti-Id, Ab2 β recognizes an idiotypic determinant within the antigen combining site and bears a structural resemblance to the original antigen (Bona and Kohler 1984). The internal image Ab2 β antibodies have been shown to induce specific immune responses to hepatitis B surface antigen in chimpanzees (Kennedy et al. 1986) and as a vaccine candidate for the control of bluetongue infection by functionally mimicking VP7 antigen of bluetongue virus (Zhou and Lin 1997). We had earlier demonstrated that an internal image Ab2 against Rinderpest virus/Peste des petits Ruminants virus H/HN protein elicits virus neutralizing antibodies as well as T cell responses in the mouse model (Vani et al. 2007a). Further, using the mouse model, DNA encoding VH region of Ab2 elicited both neutralizing antibody and T cell responses, in the complete absence of viral antigen and these responses were long lasting (Vani et al. 2007b). * M. S. Shaila shaila@mcbl.iisc.ernet.in

Infection of the skin or throat by Streptococcus dysgalactiae subspecies equisimilis (SDSE) may result in a number of human diseases. To understand mechanisms that give rise to new genetic variants in this species, we used multi-locus sequence typing (MLST) to characterise relationships in the SDSE population from India, a country where streptococcal disease is endemic. The study revealed Indian SDSE isolates have sequence types (STs) predominantly different to those reported from other regions of the world. Emm-ST combinations in India are also largely unique. Split decomposition analysis, the presence of emm-types in unrelated clonal complexes, and analysis of phylogenetic trees based on concatenated sequences all reveal an extensive history of recombination within the population. The ratio of recombination to mutation (r/m) events (11:1) and per site r/m ratio (41:1) in this population is twice as high as reported for SDSE from non-endemic regions. Recombination involving the emm-gene is also more frequent than recombination involving housekeeping genes, consistent with diversification of M proteins offering selective advantages to the pathogen. Our data demonstrate that genetic recombination in endemic regions is more frequent than non-endemic regions, and gives rise to novel local SDSE variants, some of which may have increased fitness or pathogenic potential., Journal Article, Research Support, Non-U.S. Gov't, SCOPUS: ar.j, info:eu-repo/semantics/published

Rinderpest virus belongs to the family of Paramyxoviridae, consisting of non-segmented negative sense RNA viruses. Viral transcription and replication are carried out by the RNA dependent RNA polymerase L protein which functions together with P protein as L-P complex. The exact events triggering the polymerase complex from transcription to replication function is poorly understood. In the present work, an in vitro transcription system has been described with partially purified L-P complex expressed in insect cells and viral genomic RNA. The relative abundance of each species of mRNA synthesized in vitro decreased from the 3' end of the genome to the 5' end similar to their abundance in virus infected cells. Recombinant L-P complex was unable to synthesize leader RNA suggesting the initiation of transcription from gene start site and not at the 3' end of the genome.

$CD8^+ T$ cells are thought to play an important role in protective immunity against tuberculosis. We report the identification of three peptides derived from Rv1818c, Rv3812 and Rv3018c proteins of Mycobacterium tuberculosis that bound to HLA-A*0201 molecules and their ability to induce in vitro T-cell response in peripheral blood lymphocytes from HLA-A*0201-positive healthy individuals (PPD+) and patients with TB. The peptide-specific cytotoxic T lymphocytes (CTL) generated were capable of recognizing peptide pulsed targets. Three 9-mer peptides bound with high affinity to HLA-A*0201 and displayed low dissociation rates of the bound peptide from HLA. Epitope-specific recognition was demonstrated by the release of perforin and γ-interferon. Overall, our results demonstrate the presence of HLA class I-restricted $CD8^+$ CTL against proteins from PE and PPE proteins of M. tuberculosis and identify epitopes that are strongly recognized by HLA-A*0201-restricted $CD8^+ T$ cells in humans. These epitopes thus represent potential subunit components for the design of vaccines against tuberculosis.

Three proteins of PE and PPE families of Mycobacterium tuberculosis were evaluated for their ability to induce T cell responses in mice. To enhance immunity induced by protein immunization, we tested the efficacy of adjuvant Ribi (monophosphoryl lipid A+TDM), along with three proteins of the PE/PPE family. Balb/c mice were subcutaneously injected with recombinant proteins, encoded by Rv1818c, Rv3018c and Rv3812 genes of M. tuberculosis H37Rv, formulated with Ribi or IFA for comparative study. Sera from mice immunized with Ribi revealed an increase in the specific immunoglobulin G titers by twofold against Ribi than in mice immunized with IFA. Ribi also elicited stronger delayed-type hypersensitivity and cytotoxic T-lymphocyte activity against the recombinant proteins when compared with IFA. Antigen specific IgG subclass analysis showed that Ribi tends to facilitate IgG2a production, suggesting enhancement of predominant Th1 response which in turn may facilitate increased production of protective IFN-\gamma. Furthermore, Ribi preparation increased the number of T cells secreting IFN-\gamma. These results indicate that Ribi acts as an effective adjuvant for immune response to antigens of M. tuberculosis. For the first time, we demonstrate that Rv3018c, Rv1818c and Rv3812 proteins of PE/PPE family are T cell antigens with vaccine potential.

A highly sensitive and specific reverse transcription polymerase chain reaction enzyme linked immunosorbent assay (RT-PCR-ELISA) was developed for the objective detection of nucleoprotein (N) gene of peste des petits ruminants (PPR) virus from field outbreaks or experimentally infected sheep. Two primers (IndF and Np4) and one probe (Sp3) available or designed for the amplification/probing of the 'N' gene of PPR virus, were chosen for labeling and use in RT-PCR-ELISA based on highest analytical sensitivity of detection of infective virus or N-gene containing recombinant plasmid, higher nucleotide homology at the primer binding sites of the 'N' gene sequences available and the ability to amplify PPR viral genome from different sources of samples. RT-PCR was performed with unlabeled IndF and Np4 digoxigenin labeled primers followed by a microplate hybridization probe reaction with biotin labeled Sp3 probe. RT-PCR-ELISA was found to be 10-fold more sensitive than the conventional RT-PCR followed by agarose gel based detection of PCR product. Based on the Mean (mean +/- 3S.D.) optical density (OD) values of 47 RT-PCR negative samples, OD values above 0.306 were considered positive in RT-PCR-ELISA. A total of 82 oculo-nasal swabs and tissue samples from suspected PPR cases were analyzed by RT-PCR and RT-PCR-ELISA, which revealed 54.87 and 58.54% positivity, respectively. From an experimentally infected sheep, both RT-PCR and RT-PCR-ELISA could detect the virus from 6 days post-infection up to 9 days in oculo-nasal swabs. On post-mortem, PPR viral genome was detected in spleen, lymph node, lung, heart and liver. The correlation co-efficient between RT-PCR-ELISA OD values and either TCID50 of virus or molecules of DNA was 0.622 and 0.657, respectively. The advantages of RT-PCR-ELISA over the conventional agarose gel based detection of RT-PCR products are discussed. (c) 2006 Elsevier B.V. All rights reserved.

Immunological memory is characterized by a quick and enhanced immune response after re-exposure to the same antigen. To explain the mechanism involved in generation and maintenance of immunological memory, we had earlier proposed a hypothesis involving the relay of memory by idiotypic and anti-idiotypic B cells. The peptidomimic present in the hypervariable region of anti-idiotypic antibody was hypothesized to carry forward immunological memory. In the present work, we provide evidence supporting a role for the anti-idiotypic antibody in eliciting antigen-specific B-cell and T-cell responses. Employing the idiotypic monoclonal antibody $(Ab_1)$ specific for haemagglutinin (H) protein of rinderpest virus, $Ab_{2\beta}$ was generated, which possesses an internal image of the H protein in the region between amino acids 527 and 556. We demonstrate that antigen-specific memory is perpetuated by immunization with $Ab_2$, as shown by maintenance of antigen-specific T-cell responses upon restimulation in vitro of $Ab_2$ immune splenocytes by antigen-presenting cells expressing H protein or pulsed with H-protein-derived peptides. We have also shown that boosting with antigen-specific anti-idiotypic B cells generates a memory response in antigen-primed mice. Evidence has been provided for the existence of an antigen-specific B-cell idiotypic network in the body that supports the perpetuation of immunological memory as proposed in the relay hypothesis.

Immunological memory is characterized by quick and enhanced immune response after re-exposure to the same antigen. In the present work, we have shown that anti-idiotypic T cells are generated in mice after immunization with idiotypic T cell clone or polyclonal T cell specific for nucleocapsid protein of Rinderpest virus. Further, we have shown that N specific idiotypic T cell receptors from apoptotic idiotypic T cells are processed by bone marrow derived dendritic cells and presented to cognate anti-idiotypic T cells. Evidence has been provided for the existence of antigen specific T cell idiotypic network in the body. Boosting with antigen specific anti-idiotypic T cells generates memory response in the antigen-primed mice.

$CD8^+ T$ cells are activated by the presentation of antigenic peptide through MHC class I molecules. Newly synthesized proteins formed as defective ribosomal products (DRiPs) can act as a major source of antigenic peptides for MHC class I presentation pathway. Majority of these peptides are generated from the intracellular degradation of self antigens. In the present study, we have shown that newly synthesized T cell receptor (TCR) beta chains formed as DRiPs in T cells are ubiquitinated and degraded by the proteasomes. These TCR-DRiPs are processed and presented by activated T cells to cognate anti-idiotypic $CD8^+ T$ cells. Presentation of TCR idiopeptide (peptide derived from the variable region of idiotypic TCR) by activated T cells leads to Bcl-2 expression and cytokine secretion by anti-idiotypic $CD8^+ T$ cells. Presentation of intracellular antigen by T cells may have important implications in immunoregulation, control of lymphotropic virus infection and autoimmune diseases.

A pivotal step in the initiation of T-cell immunity is the presentation of antigenic peptides by major histocompatibility complex (MHC) expressed on antigen presenting cells. The expression of MHC class II molecules by mouse T-cells has not been shown unequivocally. In the present work, we demonstrate that activated mouse T-cells synthesize MHC class II molecules de novo and express them on their surface. Further, we have demonstrated that in vitro activated T-cells take up extra-cellular soluble nucleocapsid protein of a morbillivirus. The internalized antigen goes to antigen processing compartment as shown by co-localization of antigen and LAMP-1 using confocal microscopy. We show that activated T-cells express H2M, a chaperone molecule known to have a role in antigen presentation. Further, we demonstrate that activated T-cells process and present internalized extra-cellular antigen to primed T-cells as shown by IL-2 secretion and in vitro proliferation. The presentation of antigen by T-cells may have implications in immuno-regulation, control of infection by lymphotropic viruses and maintenance of immunological memory.

The negative sense genome RNA of Rinderpest virus, a Paramyxoviridae, is encapsidated with the nucleocapsid protein N and serves as a template for the viral RNA dependent RNA polymerase for transcription and replication. The viral RNA polymerase consists of the large protein L and the phosphoprotein P functioning as the P–L complex. We provide in this report, evidences for specific binding of P protein of Rinderpest virus to the plus sense leader RNA depending on its phosphorylation status. We have also demonstrated that P protein is released from the le RNA:P protein complex upon phosphorylation in vitro. Finally, we have identified that the C-terminal 358–389 amino acid residues of P protein is involved in le RNA binding. The leader RNA binding may signify a hitherto unidentified role for P protein in the viral RNA synthesis. Moreover, our results indicate a possible role for P protein in the transcription-replication switch through leader RNA binding.

The RNA dependant RNA polymerase of negative sense RNA viruses is composed of two subunits - the Large protein ( L) and the Phosphoprotein ( P). These two proteins have to form a complex in order to carry out genome transcription and replication. Employing the baculovirus expression system, we demonstrate here, the specific in vivo interaction between the L and P proteins of Rinderpest virus and also the stabilization of L protein when it is present as L+P complex. The regions on either protein involved in such interaction has been studied using the yeast two-hybrid system which indicates that the P binding region resides within the amino terminal 380 amino acid residues of L protein. The L binding region on P protein has been mapped to lie within 347 - 490 amino acids.

The RNA dependent RNA polymerase of Rinderpest virus consists of two subunits—the large protein (L) and the phosphoprotein (P), where L is thought to be responsible for the catalytic activities in association with P protein which plays multiple roles in transcription and replication. The nucleocapsid protein (N) is necessary for encapsidation of genomic RNA, which is required as N–P complex. To understand the different steps of transcription and replication as well as the roles played by the three proteins, an in vitro reconstitution system for RNA synthesis is necessary which is not available for any morbillivirus. We describe here, an in vitro reconstitution system for transcription and replication of Rinderpest virus utilizing a synthetic, positive sense N-RNA minigenome template, free of endogenous viral polymerase proteins and recombinant viral proteins (P + L and P + N) expressed in insect cells by recombinant baculoviruses. We show that although L–P complex is sufficient to synthesize negative sense minigenome RNA, soluble N protein is necessary for encapsidation of RNA as well as synthesis of (+) sense leader RNA and (+) sense minigenome RNA.

In recent times, newly emerging infectious diseases and their causative agents have become the focus of intense investigations by medical researchers and microbiologists, particularly virologists, since many of the causative organisms are viruses. One such disease which has created so much fear world wide is severe acute respiratory syndrome (SARS). Infection with a novel corona virus has been implicated as a possible cause of SARS.Epidemiological and laboratory investigations of SARS are ongoing. WHO estimates that SARS is fatal in around 4% of cases, usually where the person has an underlying condition such as diabetes or heart disease or has a weakened immune system. In 90% of the cases people seem to recover around after a week after being infected. This is about the same mortality rate as in other diseases such as West Nile virus infection. There are currently no antiviral drugs shown to be consistently successful in treating SARS or any corona virus infection, nor anyvaccine against SARS.

The use of transgenic plants as a production system for recombinant subunit vaccines has been considered safe and economical compared to cell culture methods. We have exploited this approach to produce rinderpest virus hemagglutinin (H) protein in transgenic tobacco as a model plant for testing the immunogenicity of plant-derived hemagglutinin protein. The transgenic nature of the plants was confirmed by molecular analysis such as gene specific PCR and Southern hybridization using full-length H gene as a probe. The Mendelian pattern of inheritance of the transgene has been demonstrated in T1 generation. The transgenic plants express the H protein of molecular weight 72 kDa. The plant derived H protein is antigenically authentic as revealed by reactivity with H-specific antibodies as well as convalescent sera. The induction of immune response was tested in mice after intraperitoneal immunization with plant-derived H. High titers of antibodies were induced which were H-specific and they neutralized the infectivity of rinderpest virus.

A competitive enzyme-linked immunosorbent assay (C-ELISA) which detects antibodies unique to rinderpest virus (RPV) has been developed. This test can differentiate antibodies against RPV and those against peste des petits ruminants virus. The recombinant RPV hemagglutinin (H)-protein C-ELISA (recH C-ELISA) is based on the ability of a well-characterized monoclonal antibody (MAb) produced with the soluble, secreted form of the H protein (Sec H protein) of RPV made in a baculovirus expression system to compete with the binding of RPV antibodies in the serum of vaccinated or infected, recovered animals to the Sec H protein. The B-cell epitope recognized by the MAb corresponds to amino acids 575 to 583 on the H protein, which is not present on the antigenically closely related peste des petits ruminants virus hemagglutinin-neuraminidase protein. Initially, a positive-negative threshold cutoff value for percent inhibition of 34 was established with 500 known RPV-negative serum samples. The recH C-ELISA was developed with the enzyme immunoassay software of a commercial RPV C-ELISA kit. Comparative analysis of the test results for 700 serum samples obtained with the commercial kit gave a sensitivity of 112.4% and a specificity of 72.4%. Variations in percent inhibition values were observed for the two assay systems. These variations may have been due to the undefined amount of antigen present in the commercial kit as well as the use of a different MAb. The recH C-ELISA detected more positive serum samples compared to the number detected by the commercial kit, with the results confirmed by a virus neutralization test. Thus, recH C-ELISA is a sensitive tool for RPV serosurveillance in disease eradication programs.

We describe the properties of the two heptad repeats (HR1 and HR2) of the Peste des petits ruminants virus (PPRV) fusion protein (F) to obtain insights into the mechanism by which these repeats influence PPRV-mediated cell fusion. Both HR1 and HR2 inhibit PPRV-mediated syncytia formation in Vero cells in vitro. Of these, HR2 was found to be more effective than HR1. We studied the mechanism of fusion inhibition by these two repeats by using various biophysical and biochemical methods either separately or together. CD spectral analysis of these repeats revealed that the alpha-helical content of HR1 and HR2 when used together is higher than that of their simulated spectrum in the mixture, suggesting the formation of a highly structured complex by these repeats. Protease protection assays confirmed that such a complex is highly stable. Electrospray mass spectrometry of protease-digested products of the HR1-HR2 complex showed protection of fragments corresponding to both HR1 and HR2 sequences involved in complex formation. By employing size-exclusion chromatography and chemical cross-linking experiments, we show that three units each of HR1 and HR2 form a complex in which HR1 is a trimer and HR2 is a monomer. Homology-based three-dimensional modeling of this complex showed that HR1 and HR2 together form a six-helix and trimeric coiled-coil bundle. In this model, the HRI trimer forms the core whereas HR2, while interacting with HR1 in an antiparallel orientation, forms a two-stranded coiled-coil structure and lies at the periphery of the structure. These results are discussed in the context of a common fusion mechanism among paramyxoviruses.

To study the process of membrane fusion in Morbilliviruses, the fusion (F) glycoproteins of Peste des petits ruminants virus (PPRV) and Rinderpest virus (RPV) were expressed transiently in mammalian cells. The recombinant F proteins were found to be localized at the surface of transfected cells. The fusion activity, as evident from cell fusion assays and lysis of chicken erythrocytes, documented that transiently expressed PPRV F glycoprotein induces cell fusion in the absence of homotypic hemagglutinin–neuraminidase (HN) attachment glycoprotein. The coexpression of homotypic HN increased the extent of fusion by twofold, while the efficiency of fusion was found to be substantially enhanced. In contrast, in RPV F-expressing cells, fusion was detected only when homotypic hemagglutinin (H) or heterotypic HN protein was coexpressed. This differs from the strict type-specific requirement for the attachment protein as in the fusion process of most of the paramyxoviruses. Further, we demonstrate by fluorescence transfer experiments that while PPRV F brings about both hemifusion and complete fusion on its own, RPV F induces only hemifusion while it brings about complete fusion in the presence of homotypic or heterotypic attachment protein.

Idiotypic determinants of immunoglobulin molecules can evoke both $CD4^+$ and $CD8^+$ T responses and exist not only as the integral components of a bona fide antigen binding receptor but also as distinct molecular entities in the processed forms on the cell surface of B lymphocytes. The present work provides experimental evidence for the concept that regulation of memory B cell populations can be achieved through the presentation of idiotypic and anti-idiotypic determinants to helper and cytotoxic cell (1). The potential of B cells to present antigens to helper and cytotoxic T cells through class II and class I MHC suggests a mechanism by which both B and T cell homeostasis can be maintained. We provide evidence for the generation of idiotype- and antigen-specific Th and Tc cells upon immunization of syngenic mice with antigen or idiotypic antibody (Ab1) or anti-idiotypic antibody (Ab2). The selective activation and proliferation of the antigen-specific Th and Tc cells mediated by idiotypic stimulation observed in these experiments suggests a B-cell-driven mechanism for the maintenance of antigen-specific T cell memory in the absence of antigenic stimulation, under certain conditions.

The fusion glycoprotein (F protein) of paramyxoviruses plays a vital role in virus-induced cytopathology. To explore the role of the F protein in peste des petits ruminants virus (PPRV)- induced cytopathology, the F protein of PPRV was purified by immunoaffinity chromatography. The purified F protein, when incubated with chicken erythrocytes, caused lysis suggesting that PPRV F protein is a hemolysin. Furthermore, the hemolysis can be inhibited by hyperimmune serum against F protein. The virus-induced cell fusion (syncytia) was also inhibited by the hyperimmune serum against the F protein. In summary, these results indicate that the purified PPRV F protein is biologically active and is involved in virus-induced hemolysis, cell-fusion and the initiation of infection.

Genetic transformation systems have been established for Brassica nigra (cv. IC 257) by using an Agrobacterium binary vector as well as by direct DNA uptake of a plasmid vector. Both the type of vectors carried nptII gene and gus gene. For Agrobacterium mediated transformation, hypocotyl tissue explants were used, and up to 33% of the explants produced calli on selection medium. All of these expressed B-glucuronidase gene on histochemical staining. Protoplasts isolated from hypocotyl tissues of seedlings could be transformed with a plasmid vector by FEG mediated uptake of vector DNA. A number of fertile kanamycin resistant plants were obtained using both the methods, and their transformed nature was confirmed by Southern blot analysis and histochemical staining for GUS. Backcrossed and selfed progenies of these transformed plants showed the presence of npt and gus genes.

The construction and characterization of two genome-specific recombinant DNA clones from B. nigra are described. Southern analysis showed that the two clones belong to a dispersed repeat family. They differ from each other in their length, distribution and sequence, though the average GC content is nearly the same (45%). These B genome-specific repeats have been used to analyse the phylogenetic relationships between cultivated and wild species of the family Brassicaceae.

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.