Progress 10/01/09 to 09/30/10
Outputs
Progress Report Objectives (from AD-416) This proposal presents a broad community plan to begin to systematically address the immunological reagent gap for the U.S. veterinary immunology research community including for the following groups: ruminants (concentrating on cattle), swine, poultry (primarily chickens with some evaluation of reagents on turkey cells), horses, and aquaculture species (concentrating on channel catfish and trout) with a goal of 20 reagents per species group. The reagents will include bioactive recombinant cytokines and chemokine proteins, expressed using mammalian, yeast, and bacterial systems, as well as antibodies to them and their receptors as well as antibodies to immunoglobulin isotypes, T cell receptors and other CD molecules. The goal is to produce antibodies that function in ELISA and ELISpot assays, for intracellular staining, for blocking function and signaling, and that are useful in flow cytometric applications as well as in fixed tissue sections. Antibodies to bioactive molecules will also be tested for their ability to block the recombinant molecules as well as natural molecules if it is possible to produce cell culture systems for the later. This project represents the swine component. Approach (from AD-416) Prioritization of reagents (tools) to be produced will be established by surveying the larger scientific community and cooperating with other similar international efforts to avoid redundancy and target the most useful reagents for research scientists. Commercial companies will also be surveyed. Bioactive cytokines and chemokines and cell surface CD molecules, antibodies, and T cell receptors will be produced in expression systems in two main labs for poultry and mammals and by the two co-PDs for fish. The expression of these proteins will depend upon provision of cDNA from appropriate cells or tissues from the co-PD labs who are specialists in particular species. RT-PCR using primers based on available sequences or from homologous genes in other related species will be done to obtain the entire gene sequence. The recombinant molecules will be used to prime mice for production of monoclonal antibodies or rabbits for polyclonal antibodies in a centralized lab. After identifying reactive antibodies initially, both the recombinant molecules and the antibodies to them will be evaluated for efficacy in the species specialist labs in a variety of in vitro assays. This project will have oversight by an advisory board composed of industry partners, USDA national program directors and other international reagent- generating groups. The reagents will be provided to the scientific community by depositing in commercial cell banks and encouraging generation and distribution through commercial vendors. In this Agreement swine immune reagents were developed to probe immune responses to infectious diseases and to improve vaccine design. The U.S. Veterinary Immunological Reagents Network (VIRN) (www.vetimm.org) was established to address the dearth of immunological reagents for food animal species. ARS Researchers at Beltsville, MD, working with Kingfisher Biotech Inc. www.kingfisherbiotech.com/ have expressed a total of 16 recombinant swine immune proteins in yeast. This year 6 recombinant chemokines and cytokines (CCL2, CCL3L1, CCL4, CCL5, IFNalpha, IL-15) have been proven to be bioactive. A panel of monoclonal antibodies (mAbs) against CCL2 was produced at Cornell Univ., characterized, and a new Luminex bead assay for quantitating CCL2 was jointly developed. One mAb each reactive with IL-4R and IL-13R has been characterized; new hybridoma fusions were screened for anti-T cell specific mAb with Cornell Univ. scientists. Products generated by the VIRN will be used by animal health researchers, veterinarians, vaccine manufacturers, and other commercial sources. The U.S. Veterinary Immunological Reagents Network (VIRN) website for swine immunological reagents was expanded over the past year. ARS Researchers at Beltsville, MD, have provided all documentation and knowhow for the USDA CSREES grant supported immune toolkit or VIRN website (http://www.umass.edu/vetimm/swine/index.html) swine pages. They scanned commercial websites and publications for swine immune reagents and organized the data into easily accessible Tables of currently available reagents for swine immunology. This list served as the basis for updating the Swine priorities for developing new VIRN immune reagents and for VIRN colleagues and swine collaborators to develop plans for their production. Posted results as well as updated Protocols on the VIRN website. As a result this website enables researchers worldwide to quickly identify available immune tools for their experiments. Overall swine disease and vaccine researchers have improved tools to address pig health and vaccine design. Communications for this grant have been supported by ARS by providing conference call line access for the monthly calls that review Network plans and progress. Email and phone communication are regularly exchanged between the participating labs as specific reagents are developed and screened.
Impacts
(N/A)
Publications
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Progress 01/15/06 to 07/31/10
Outputs
Progress Report Objectives (from AD-416) This proposal presents a broad community plan to begin to systematically address the immunological reagent gap for the U.S. veterinary immunology research community including for the following groups: ruminants (concentrating on cattle), swine, poultry (primarily chickens with some evaluation of reagents on turkey cells), horses, and aquaculture species (concentrating on channel catfish and trout) with a goal of 20 reagents per species group. The reagents will include bioactive recombinant cytokines and chemokine proteins, expressed using mammalian, yeast, and bacterial systems, as well as antibodies to them and their receptors as well as antibodies to immunoglobulin isotypes, T cell receptors and other CD molecules. The goal is to produce antibodies that function in ELISA and ELISpot assays, for intracellular staining, for blocking function and signaling, and that are useful in flow cytometric applications as well as in fixed tissue sections. Antibodies to bioactive molecules will also be tested for their ability to block the recombinant molecules as well as natural molecules if it is possible to produce cell culture systems for the later. This project represents the swine component. Approach (from AD-416) Prioritization of reagents (tools) to be produced will be established by surveying the larger scientific community and cooperating with other similar international efforts to avoid redundancy and target the most useful reagents for research scientists. Commercial companies will also be surveyed. Bioactive cytokines and chemokines and cell surface CD molecules, antibodies, and T cell receptors will be produced in expression systems in two main labs for poultry and mammals and by the two co-PDs for fish. The expression of these proteins will depend upon provision of cDNA from appropriate cells or tissues from the co-PD labs who are specialists in particular species. RT-PCR using primers based on available sequences or from homologous genes in other related species will be done to obtain the entire gene sequence. The recombinant molecules will be used to prime mice for production of monoclonal antibodies or rabbits for polyclonal antibodies in a centralized lab. After identifying reactive antibodies initially, both the recombinant molecules and the antibodies to them will be evaluated for efficacy in the species specialist labs in a variety of in vitro assays. This project will have oversight by an advisory board composed of industry partners, USDA national program directors and other international reagent- generating groups. The reagents will be provided to the scientific community by depositing in commercial cell banks and encouraging generation and distribution through commercial vendors. BL-1, 9/27/06. No research was performed on this project this year. This project was only kept open for closeout and that process has been completed.
Impacts
(N/A)
Publications
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Progress 10/01/08 to 09/30/09
Outputs
Progress Report Objectives (from AD-416) This proposal presents a broad community plan to begin to systematically address the immunological reagent gap for the U.S. veterinary immunology research community including for the following groups: ruminants (concentrating on cattle), swine, poultry (primarily chickens with some evaluation of reagents on turkey cells), horses, and aquaculture species (concentrating on channel catfish and trout) with a goal of 20 reagents per species group. The reagents will include bioactive recombinant cytokines and chemokine proteins, expressed using mammalian, yeast, and bacterial systems, as well as antibodies to them and their receptors as well as antibodies to immunoglobulin isotypes, T cell receptors and other CD molecules. The goal is to produce antibodies that function in ELISA and ELISpot assays, for intracellular staining, for blocking function and signaling, and that are useful in flow cytometric applications as well as in fixed tissue sections. Antibodies to bioactive molecules will also be tested for their ability to block the recombinant molecules as well as natural molecules if it is possible to produce cell culture systems for the later. This project represents the swine component. Approach (from AD-416) Prioritization of reagents (tools) to be produced will be established by surveying the larger scientific community and cooperating with other similar international efforts to avoid redundancy and target the most useful reagents for research scientists. Commercial companies will also be surveyed. Bioactive cytokines and chemokines and cell surface CD molecules, antibodies, and T cell receptors will be produced in expression systems in two main labs for poultry and mammals and by the two collaborators for fish. The expression of these proteins will depend upon provision of cDNA from appropriate cells or tissues from the collaborator labs who are specialists in particular species. RT-PCR using primers based on available sequences or from homologous genes in other related species will be done to obtain the entire gene sequence. The recombinant molecules will be used to prime mice for production of monoclonal antibodies or rabbits for polyclonal antibodies in a centralized lab. After identifying reactive antibodies initially, both the recombinant molecules and the antibodies to them will be evaluated for efficacy in the species specialist labs in a variety of in vitro assays. This project will have oversight by an advisory board composed of industry partners, USDA national program directors and other international reagent-generating groups. The reagents will be provided to the scientific community by depositing in commercial cell banks and encouraging generation and distribution through commercial vendors. Significant Activities that Support Special Target Populations A panel of immunological reagents have been developed to improve studies on immune responses to infectious diseases and to improve swine vaccine design. This USDA CSREES grant established the U.S. Veterinary Immunological Reagents Network (VIRN) (www.vetimm.org) to address the dearth of immunological reagents. Working with Kingfisher Biotech Inc., swine immune proteins were express in yeast; 5 recombinant chemokines (CXCL10, CXCL11, CCL2, CCL4 and CCL5) have been proven to be bioactive at BARC and monoclonal antibodies (mAbs) produced to several of these and now being characterized. Separately, Cornell U. scientists used recombinant cell surface proteins to produce mAb for 2 swine cytokine receptors, IL-4R and IL-13R; their reactivity has been affirmed at BARC. Products generated by the VIRN will be used by animal health researchers, veterinarians, vaccine manufacturers, and other commercial sources. The U. S. Veterinary Immunological Reagents Network (VIRN) website for swine immunological reagents has been expanded. BARC APDL scientists have provided all documentation and knowhow for the USDA CSREES grant supported immune toolkit or VIRN website (http://www.umass. edu/vetimm/swine/index.html) swine pages. They scanned commercial websites and publications for swine immune reagents and organized the data into easily accessible Tables of currently available reagents for swine immunology. With this basis they updated the Swine priorities for new immune reagents, worked with VIRN colleagues to develop them, and posted those results as well as the techniques to assess their bioactivity. Details of Network progress in developing cloned cytokine and chemokine proteins and monoclonal antibodies to them and to cell surface proteins can now be easily monitored, and requests for reagents reviewed. As a result, researchers worldwide can quickly access lists of available commercial and planned VIRN reagents for swine immune and disease studies. This website enables swine disease and vaccine researchers to quickly identify the availability of immune tools for their experiments and thus will help to improve pig health and vaccine design.
Impacts
(N/A)
Publications
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Progress 10/01/07 to 09/30/08
Outputs
Progress Report Objectives (from AD-416) This proposal presents a broad community plan to begin to systematically address the immunological reagent gap for the U.S. veterinary immunology research community including for the following groups: ruminants (concentrating on cattle), swine, poultry (primarily chickens with some evaluation of reagents on turkey cells), horses, and aquaculture species (concentrating on channel catfish and trout) with a goal of 20 reagents per species group. The reagents will include bioactive recombinant cytokines and chemokine proteins, expressed using mammalian, yeast, and bacterial systems, as well as antibodies to them and their receptors as well as antibodies to immunoglobulin isotypes, T cell receptors and other CD molecules. The goal is to produce antibodies that function in ELISA and ELISpot assays, for intracellular staining, for blocking function and signaling, and that are useful in flow cytometric applications as well as in fixed tissue sections. Antibodies to bioactive molecules will also be tested for their ability to block the recombinant molecules as well as natural molecules if it is possible to produce cell culture systems for the later. This project represents the swine component. Approach (from AD-416) Prioritization of reagents (tools) to be produced will be established by surveying the larger scientific community and cooperating with other similar international efforts to avoid redundancy and target the most useful reagents for research scientists. Commercial companies will also be surveyed. Bioactive cytokines and chemokines and cell surface CD molecules, antibodies, and T cell receptors will be produced in expression systems in two main labs for poultry and mammals and by the two co-PDs for fish. The expression of these proteins will depend upon provision of cDNA from appropriate cells or tissues from the co-PD labs who are specialists in particular species. RT-PCR using primers based on available sequences or from homologous genes in other related species will be done to obtain the entire gene sequence. The recombinant molecules will be used to prime mice for production of monoclonal antibodies or rabbits for polyclonal antibodies in a centralized lab. After identifying reactive antibodies initially, both the recombinant molecules and the antibodies to them will be evaluated for efficacy in the species specialist labs in a variety of in vitro assays. This project will have oversight by an advisory board composed of industry partners, USDA national program directors and other international reagent- generating groups. The reagents will be provided to the scientific community by depositing in commercial cell banks and encouraging generation and distribution through commercial vendors. Significant Activities that Support Special Target Populations A major obstacle to advances in veterinary immunology and disease control is the lack of immunological reagents. This grant established the US Veterinary Immunological Reagents Network (VIRN) (www.vetimm.org) to address the dearth of immunological reagents. BARC APDL scientists provided materials to the consortium including tissue specific cDNAs, plasmids, primers, design of peptides, recombinant proteins, nucleotide and peptide sequences and associated documentation and knowhow. VIRN labs have produced full-length plasmids for 19 genes. Yeast expression vectors that encode swine immune proteins were used to express 5 recombinant chemokines and cytokines at Kingfisher Biotech. To date 2 chemokines and 1 cytokine have been tested for bioactivity at BARC APDL and will be sent to U. Massachusetts for monoclonal antibody (MAb) production. Separately Cornell U. scientists used swine cDNAs for expression of cell surface proteins to produce MAb; to date MAb for 2 swine T cell receptors and one cytokine receptor have been screened for activity. Products generated by the VIRN will be used by animal health researchers, veterinarians, vaccine manufacturers, and other commercial applications. The VIRN website has been expanded. BARC APDL scientists have provided all documentation and knowhow for the VIRN website www.vetimm.org expansion. They reviewed, and regularly updated, published data and techniques on swine immune reagents and their bioactivity. In addition, they scanned commercial websites for swine immune reagents, and organized the data into easily accessible Tables. As a result researchers worldwide can quickly access lists of available commercial reagents for swine immune and disease studies. Details of Network progress in developing cloned cytokine and chemokine proteins and monoclonal antibodies to them and to cell surface proteins can be easily monitored, and requests for reagents can be honored through the website and the Network�s commercial partner. Additionally, the website provides a forum on which researchers can suggest new priorities for future development. This website enables swine disease and vaccine researchers to quickly identify the availability of immune tools for their experiments and thus will help to improve pig health and vaccine design. These projects align with National Program 103 Component II: Genetic and biological determinants of disease susceptibility. Communications for this grant have been supported by ARS by providing support for the monthly conference calls that review Network plans and progress. An ARS Sharepoint site has just been established for the Network for more effective data exchange. Email and phone communication are regularly exchanged between the participating labs as specific reagents are developed and screened.
Impacts
(N/A)
Publications
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Progress 10/01/06 to 09/30/07
Outputs
Progress Report Objectives (from AD-416) This proposal presents a broad community plan to begin to systematically address the immunological reagent gap for the U.S. veterinary immunology research community including for the following groups: ruminants (concentrating on cattle), swine, poultry (primarily chickens with some evaluation of reagents on turkey cells), horses, and aquaculture species (concentrating on channel catfish and trout) with a goal of 20 reagents per species group. The reagents will include bioactive recombinant cytokines and chemokine proteins, expressed using mammalian, yeast, and bacterial systems, as well as antibodies to them and their receptors as well as antibodies to immunoglobulin isotypes, T cell receptors and other CD molecules. The goal is to produce antibodies that function in ELISA and ELISpot assays, for intracellular staining, for blocking function and signaling, and that are useful in flow cytometric applications as well as in fixed tissue sections. Antibodies to bioactive molecules will also be tested for their ability to block the recombinant molecules as well as natural molecules if it is possible to produce cell culture systems for the later. This project represents the swine component. Approach (from AD-416) Prioritization of reagents (tools) to be produced will be established by surveying the larger scientific community and cooperating with other similar international efforts to avoid redundancy and target the most useful reagents for research scientists. Commercial companies will also be surveyed. Bioactive cytokines and chemokines and cell surface CD molecules, antibodies, and T cell receptors will be produced in expression systems in two main labs for poultry and mammals and by the two co-PDs for fish. The expression of these proteins will depend upon provision of cDNA from appropriate cells or tissues from the co-PD labs who are specialists in particular species. RT-PCR using primers based on available sequences or from homologous genes in other related species will be done to obtain the entire gene sequence. The recombinant molecules will be used to prime mice for production of monoclonal antibodies or rabbits for polyclonal antibodies in a centralized lab. After identifying reactive antibodies initially, both the recombinant molecules and the antibodies to them will be evaluated for efficacy in the species specialist labs in a variety of in vitro assays. This project will have oversight by an advisory board composed of industry partners, USDA national program directors and other international reagent- generating groups. The reagents will be provided to the scientific community by depositing in commercial cell banks and encouraging generation and distribution through commercial vendors. Significant Activities that Support Special Target Populations This report documents research conducted under a Reimbursable Agreement between ARS and the Univ. of Massachusetts through USDA CSREES grant to Univ. Massachusetts, Amherst for the Veterinary Immunological Reagents Network with the USDA ARS BARC APDL scientist serving as the Chair for swine reagents. Additional details of research can be found in the report for the parent project 1265-32000-079-00D, "Strategies to Control Swine Parasites Affecting Food Safety." A major obstacle to advances in veterinary immunology and disease control is the lack of sufficient immunological reagents. This USDA CSREES was funded in 2006 through a USDA CSREES grant to Univ. Massachusetts, Amherst, to establish the Veterinary Immunological Reagents Network. The Network was established to address the immunological reagent gap for the US veterinary immunology research community. This is 4 year multi-species effort supports cloning and expression of immunological reagents specific for ruminants, swine, poultry, equine and aquaculture species. The USDA ARS BARC APDL scientist serves as the Chair for swine reagents. The emphases for swine is on developing and characterizing bioactive immune proteins, cloned cytokines (interferons and interleukins) and chemokine proteins, as well as monoclonal antibodies (mAbs) to these proteins and their receptors. The prioritiy list is maintained on the network website http://www.umass. edu/vetimm/swine/index.html. Additional mAb will be characterized for toll-like receptor (TLR) proteins or produced to swine cell subset proteins, the CD antigens, and the T cell receptors. Reagents developed with this grant will be used for disease and vaccine research by veterinary immunologists, pathologists and microbiologists. National Program 103 Component 2: Genetic and biological determinants of disease susceptibility The ADODR for this grant helped to establish, and regularly participate in, monthly conference calls that review plans and progress for reagent development under this grant. Rregular email contact with the participating labs as they develop reagents were also exchanged.
Impacts
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Publications
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Progress 10/01/05 to 09/30/06
Outputs
Progress Report 4d Progress report. This report serves to document research conducted under a Reimbursable Agreement through USDA CSREES grant to Univ. Massachusetts, Amherst for the Veterinary Immunological Reagents Network with the USDA ARS BARC APDL scientist serving as the Chair for swine reagents. Additional details of research can be found in the report from the parent project 1265-32000- 079-00D: Strategies to Control Swine Parasites Affecting Food Safety. A major obstacle to advances in veterinary immunology and disease control is the lack of sufficient immunological reagents. This USDA CSREES for a Veterinary Immunological Reagents Network, was funded in 2006 through a USDA CSREES grant to Univ. Massachusetts, Amherst, to establish the Veterinary Immunological Reagents Network to begin to address this immunological reagent gap for the US veterinary immunology research community. This is a 4 year multi-species effort which will support immunological reagents
specific for ruminants, swine, poultry, equine and aquaculture species, with the USDA ARS BARC APDL scientist serving as the Chair for swine reagents. The emphases for swine will be on developing and characterizing bioactive immune proteins, cloned cytokine and chemokine proteins, as well as monoclonal antibodies (mAbs) to these proteins and their receptors. Additional mAb will be produced to swine cell subset proteins, the CD antigens, and toll-like receptor (TLR) proteins. Reagents developed with this grant will be used for disease and vaccine research by veterinary immunologists, pathologists and microbiologists.
Impacts
(N/A)
Publications
- Baldwin, C., Hansen, J.D., Horohov, D., Labresh, J., Lillehoj, H.S., Lunney, J.K., Miller, N.W., Wilson, M., Bengten, E., Chinchar, G., Wagner, B. 2006. The U.S. veterinary immune reagents network. In: Proceedings of International Veterinary Vaccines and Diagnostics Conference, June 25-29, 2006, Oslo, Norway. p. 34.
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