MONOCLONAL-ANTIBODIES TO DISTINCTIVE EPITOPES ON THE ALPHA AND BETA SUBUNITS OF THE FIBRONECTIN RECEPTOR
MONOCLONAL-ANTIBODIES TO DISTINCTIVE EPITOPES ON THE ALPHA AND BETA SUBUNITS OF THE FIBRONECTIN RECEPTOR
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DOI：
10.1016/0014-4827(88)90464-8
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发表时间：
1988-08-01
影响因子：
3.7
通讯作者：
JULIANO, RL
中科院分区：
文献类型：
--
作者：
BROWN, PJ;JULIANO, RL
已经开发了单克隆抗体（MAb），其可以识别对α-半乳糖苷酶或α-半乳糖苷酶独特的表位。或β纤连蛋白受体（FnR）亚基。MAb 11B 4和7A 8免疫印迹α-葡聚糖。从中国仓鼠卵巢（CHO）细胞中纯化的FnR亚基，或在还原或非还原条件下纯化的人和啮齿动物来源细胞的非离子去污剂提取物中。单克隆抗体似乎是更具反应性的亚基时，它已被还原条件下，表明表位可能被部分掩盖的构象赋予二硫键。第二组MAb，7 E2和7 F9，针对β-内酰胺酶上的表位。在一些实施方案中，所述单克隆抗体是构象上依赖于二硫键的亚基，因为亚基的还原导致与两种MAb的反应性丧失。CHO细胞的非离子去污剂提取物的进一步的7 E2/7 F9免疫印迹，在非还原条件下进行，揭示了第三条带（90-kDa）的存在，其在免疫学上与β-内酰胺酶相关。亚基，其在完整细胞中未被125 I表面标记，并且不与α亚基共纯化。和β通过使用MAb PB 1对FnR进行免疫亲和纯化来分离FnR亚基。未发现90-kDa组分与通过粗细胞分级制备的质膜组分相关，但在含有细胞核和细胞内膜的低速颗粒中丰富。这一发现表明，90-kDa组分是β的前体。亚单位最后，7 E2/7 F9的表位对于CHO细胞是独特的，因为与其他细胞类型的交叉反应性不能通过免疫印迹或免疫沉淀来证明。
Monoclonal antibodies (MAbs) have been developed that can recognize epitopes that are unique to either the .alpha. or .beta. subunit of the fibronectin receptor (FnR). MAbs 11B4 and 7A8 immunoblot the .alpha. subunit of FnR either in purified form from Chinese hamster ovary (CHO) cells or in nonionic detergent extracts of cells of human and rodent origin electrophoresed under reducing or nonreducing conditions. The MAbs seem to be more reactive to the subunit when it has been electrophoresed under reducing conditions, suggesting that the epitope may be partially masked by the conformation conferred by disulfide bonding. A second set of MAbs, 7E2 and 7F9, is directed to an epitope on the .beta. subunit that is conformationally dependent upon disulfide bonding, as reduction of the subunit leads to loss of reactivity with both MAbs. Further 7E2/7F9 immunoblots of nonionic detergent extracts of CHO cells, run under nonreducing conditions, reveal the presence of a third band (90-kDa), immunologically related to the .beta. subunit, which is not surface-labeled with 125I in intact cells and which does not copurify with the .alpha. and .beta. subunits isolated by immunoaffinity purification of FnR using the MAb PB1. The 90-kDa component is not found associated with a plasma membrane fraction prepared by crude cell fractionation, but is abundant in a low-speed pellet containing nuclei and intracellular membranes. This finding suggests that the 90-kDa component is a precursor to the .beta. subunit. Finally, the epitope of 7E2/7F9 is unique to CHO cells, as cross-reactivity to other cell types cannot be demonstrated by either immunoblotting or immunoprecipitation.