Ghebrehiwet B, Lu P D, Zhang W, Lim B L, Eggleton P, Leigh L E, Reid K B, Peerschke E I
Department of Medicine, State University of New York, Stony Brook 11794, USA.
Hybridoma. 1996 Oct;15(5):333-42. doi: 10.1089/hyb.1996.15.333.
A membrane protein (33 kDa) that binds to the globular "heads" of C1q (gC1q-R) has been recently described. The full length cDNA encoding gC1q-R has been cloned, expressed in E. coli and using the purified recombinant protein (rgC1q-R) as an immunogen, a panel of IgG monoclonal antibodies (MAb) has been produced by fusion of spleen cells from hyperimmunized BALB/c mice with NSO mouse myeloma partners. From this fusion, 60 anti-gC1q-R hybridomas were selected and evaluated for their ability to (1) discriminate between the mature form (MF) of gC1q-R (residues 74-282) and a truncated form (TF) lacking residues 74-95, which contains a major C1q binding site, (2) recognize two functionally defined synthetic peptides derived from the NH2-(XN18) and COOH-(XC15) terminus of gC1q-R, and (3) bind to microtiter well fixed intact Raji cells. Several clones were identified: MAbs 46.23 and 60.11 (IgG1 kappa), reacted strongly with ELISA plate-fixed intact Raji and K562 cells, MF, and the XN18 peptide, but had poor or no reactivity with TF; MAbs 74.5.2 > 25.15 (IgG1 kappa) recognized both MF and TF and are directed against epitopes in the XC15 peptide that contains a binding site for high-molecular-weight kininogen and Factor XII.
最近发现了一种与C1q球状“头部”（gC1q-R）结合的膜蛋白（33 kDa）。编码gC1q-R的全长cDNA已被克隆，并在大肠杆菌中表达。利用纯化的重组蛋白（rgC1q-R）作为免疫原，通过将超免疫BALB/c小鼠的脾细胞与NSO小鼠骨髓瘤细胞系融合，制备了一组IgG单克隆抗体（MAb）。从该融合产物中，筛选出60个抗gC1q-R杂交瘤，并评估它们对以下方面的能力：（1）区分gC1q-R的成熟形式（MF，残基74 - 282）和缺少残基74 - 95的截短形式（TF），后者包含一个主要的C1q结合位点；（2）识别源自gC1q-R NH2端（XN18）和COOH端（XC15）的两个功能明确的合成肽；（3）结合微孔板固定的完整Raji细胞。鉴定出了几个克隆：单克隆抗体46.23和60.11（IgG1 κ）与ELISA板固定的完整Raji和K562细胞、MF以及XN18肽强烈反应，但与TF反应较弱或无反应；单克隆抗体74.5.2 > 25.15（IgG1 κ）识别MF和TF，并且针对XC15肽中的表位，该表位包含高分子量激肽原和因子XII结合位点。
