Stable expression of the extracellular domains of rabbit recombinant CD5: development and characterization of polyclonal and monoclonal antibodies.
Previous studies in our laboratory suggested that there was positive selection of B cells during early development in the appendix of normal and V(H) mutant (ali/ali) rabbits. Preferential expansion and survival of B lymphocytes was affected by the Ig V(H) frameworks 1 and 3 sequences expressed on the cell surface. We demonstrated a specific interaction between rabbit CD5 and the V region of rabbit heavy chains and suggested that CD5 is a potential selecting ligand for B-cell surface immunoglobulin framework region sequences. To further investigate the role of CD5 in rabbit B-cell selection and survival we prepared recombinant constructs and obtained stable expression of the three scavenger receptor cysteine-rich (SRCR) extracellular domains of rabbit CD5. Here we describe the production and purification of this expressed recombinant CD5 protein, polyclonal antibody obtained by immunization of a goat and initial production and characterization of specific mAbs against peptides selected from each sequenced SRCR domain.
Pospisil
Richard
R
Laboratory of Immunology, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Building 10, Room llN3ll, 10 Center Drive-MSC 1892, Bethesda, MD 20892-1892, USA.
Obiakor
Harold
H
Newman
Barbara A
BA
Alexander
Cornelius
C
Mage
Rose G
RG
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis* / Q000276:immunology; D001065:Appendix / Q000276:immunology; D001402:B-Lymphocytes / Q000276:immunology*; D001483:Base Sequence; D015153:Blotting, Western; D018956:CD5 Antigens / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology; D007135:Immunoglobulin Variable Region / Q000235:genetics / Q000276:immunology*; D007150:Immunohistochemistry; D008969:Molecular Sequence Data; D010957:Plasmids; D012313:RNA / Q000737:chemistry / Q000235:genetics; D011817:Rabbits / Q000276:immunology*; D011971:Receptors, Immunologic / Q000276:immunology; D051116:Receptors, Scavenger; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000662:veterinary; D016415:Sequence Alignment; D013154:Spleen / Q000276:immunology; D014170:Transformation, Genetic
D000911:Antibodies, Monoclonal; D018956:CD5 Antigens; D007135:Immunoglobulin Variable Region; D011971:Receptors, Immunologic; D051116:Receptors, Scavenger; D011994:Recombinant Proteins; D012313:RNA