Banks L, Matlashewski G, Crawford L
Eur J Biochem. 1986 Sep 15;159(3):529-34. doi: 10.1111/j.1432-1033.1986.tb09919.x.
The isolation and construction of a complete human p53 cDNA and subsequent expression in monkey cells is described. A set of new anti-(human p53) monoclonal antibodies has also been obtained and used to show the expression of the human p53 cDNA in cos-l cells. These antibodies enable the specific detection of human p53, which is synthesised in the presence of p53 from other species. Fusion proteins of p53 with beta-galactosidase were used firstly as antigen and secondly, in conjunction with competition assays, to localise the determinants recognized by the antibodies. At least two previously unrecognized epitopes are involved and two of the antibodies are human-p53-specific. The epitopes are denaturation-resistant and the antibodies are, therefore, valuable for immunoblotting as well as immunoprecipitation and enzyme-linked immunoassay. Transfection of plasmids containing complete human p53 cDNA into monkey (cos-l) cells cause expression of human p53 recognized by the monoclonal antibodies. Control plasmids did not induce immunoreactive protein.
本文描述了完整人p53 cDNA的分离与构建，以及随后在猴细胞中的表达。还获得了一组新的抗（人p53）单克隆抗体，并用于检测人p53 cDNA在cos-1细胞中的表达。这些抗体能够特异性检测人p53，即使在存在其他物种p53的情况下合成的人p53也能被检测到。p53与β-半乳糖苷酶的融合蛋白首先用作抗原，其次结合竞争试验来定位抗体识别的决定簇。至少涉及两个以前未被识别的表位，其中两种抗体是人p53特异性的。这些表位具有抗变性能力，因此这些抗体对于免疫印迹、免疫沉淀和酶联免疫测定都很有价值。将含有完整人p53 cDNA的质粒转染到猴（cos-1）细胞中，可导致单克隆抗体识别的人p53表达。对照质粒未诱导免疫反应性蛋白。
