Identification of monoclonal antibodies that recognize different disulfide bonded forms of thrombospondin 1.
Thrombospondin 1 (TSP1) is a multidomain glycoprotein from platelets and cells which functions in cell-cell and cell-matrix interactions. The structure of TSP1 is regulated by sulfhydryl-disulfide interchange in the carboxy-terminal Ca2(+)-binding loops and globular domain which markedly influence its interaction with cell surface integrins and its inhibition of neutrophil enzymes. We have identified murine monoclonal antibodies that recognized different disulfide-bonded forms of TSP1, made by preparing TSP1 in buffers containing either 0.1 mM or 2 mM Ca2+. Antibody HB8432 recognizes TSP1 prepared in buffers containing either 0.1 or 2 mM Ca2+, while antibodies D4.6 and A65M recognized only TSP1 prepared in buffers containing 0.1 mM Ca2+. The antibodies recognize these different TSP1 preparations either adsorbed to plastic or extracellular matrix. Immunohistochemistry of human rheumatoid synovial tissue using HB8432 resulted in staining of numerous blood vessel walls and matrix cells, while D4.6 and A65M stained a subset of the HB8432 positive blood vessels and only occasionally stained matrix cells. These results suggested that different disulfide-bonded forms of TSP1 were being expressed in different areas of inflamed tissue.
Matthias
L J
LJ
Centre for Thrombosis and Vascular Research, School of Pathology, University of New South Wales, Sydney, Australia.
Gotis-Graham
I
I
Underwood
P A
PA
McNeil
H P
HP
Hogg
P J
PJ
D000327:Adsorption; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000918:Antibody Specificity; D000937:Antigen-Antibody Reactions; D001172:Arthritis, Rheumatoid / Q000276:immunology / Q000378:metabolism; D003094:Collagen; D003553:Cystine / Q000737:chemistry*; D004338:Drug Combinations; D000939:Epitopes / Q000737:chemistry / Q000276:immunology*; D006801:Humans; D007797:Laminin; D008562:Membrane Glycoproteins / Q000032:analysis / Q000737:chemistry / Q000276:immunology*; D051379:Mice; D010969:Plastics; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011509:Proteoglycans; D011817:Rabbits; D013583:Synovial Membrane / Q000737:chemistry; D019699:Thrombospondins
D000911:Antibodies, Monoclonal; D004338:Drug Combinations; D000939:Epitopes; D007797:Laminin; D008562:Membrane Glycoproteins; D010969:Plastics; D011509:Proteoglycans; D019699:Thrombospondins; C059485:matrigel; D003553:Cystine; D003094:Collagen