Monoclonal antibodies directed against epitopes within the core protein structure of the large aggregating proteoglycan (aggrecan) from the swarm rat chondrosarcoma.
The core protein of the large hyaline cartilage proteoglycan, aggrecan, is composed of six distinct domains: globular 1 (G1), interglobular, globular 2 (G2), keratan sulfate attachment, chondroitin sulfate (CS) attachment, and globular 3 (G3). Monoclonal antibodies that recognize epitopes in these domains were raised against Swarm rat chondrosarcoma aggrecan that was either denatured through reduction and alkylation or partially deglycosylated through chondroitinase ABC digestion or alkali elimination, the latter with or without sulfite addition. Monoclonal antibodies were further characterized for reactivity to purified aggrecan substructures including rat chondrosarcoma G1 and CS attachment domains, a recombinant rat chondrosarcoma G3 domain fusion protein, bovine articular cartilage G2 domain, and rat chondrosarcoma link protein (LP). Biochemical characterization of the specificities of these monoclonal antibodies indicated that one (1C6) recognized an epitope shared by both the G1 and the G2 domains; one (5C4) recognized an epitope shared by both LP and the G1 domain; one (7D1) recognized an epitope shared by both the G1 and the CS attachment domains; two (14A1 and 15B2) recognized epitopes in the CS attachment domain; one (14B4) recognized an epitope in the G3 domain; and one (13D1) recognized a ubiquitous epitope shared by the G1, G2, G3, and CS attachment domains of aggrecan and also LP. Collectively the specificities of these antibodies confirm the occurrence of multiple repeated epitopes (both carbohydrate and protein in nature) throughout the different domain structures of aggrecan. These antibodies have been proven to be useful for identifying aggrecan-like molecules in several connective tissues other than cartilage.
Calabro
A
A
Bone Research Branch, National Institute of Dental Research, National Institutes of Health, Bethesda, Maryland 20892.
Hascall
V C
VC
Caterson
B
B
D053674:Aggrecans; D000818:Animals; D000911:Antibodies, Monoclonal; D000917:Antibody Formation; D002358:Cartilage, Articular / Q000737:chemistry; D002417:Cattle; D011508:Chondroitin Sulfate Proteoglycans / Q000032:analysis / Q000737:chemistry / Q000276:immunology*; D002813:Chondrosarcoma / Q000737:chemistry*; D000939:Epitopes / Q000032:analysis*; D016326:Extracellular Matrix Proteins; D005260:Female; D037181:Lectins, C-Type; D051379:Mice; D008807:Mice, Inbred BALB C / Q000276:immunology; D008961:Models, Structural; D011487:Protein Conformation; D011509:Proteoglycans / Q000032:analysis / Q000737:chemistry / Q000276:immunology*; D051381:Rats
C508320:Acan protein, mouse; C508321:Acan protein, rat; D053674:Aggrecans; D000911:Antibodies, Monoclonal; D011508:Chondroitin Sulfate Proteoglycans; D000939:Epitopes; D016326:Extracellular Matrix Proteins; D037181:Lectins, C-Type; D011509:Proteoglycans