Margulis B A, Nacharov P V, Tsvetkova O I, Welsh M, Kinev A V
Institute of Cytology, USSR Academy of Sciences, Leningrad.
Electrophoresis. 1991 Sep;12(9):670-3. doi: 10.1002/elps.1150120913.
The hsp70 family of major stress proteins is composed of several different members exhibiting similar structural and functional properties. In order to obtain an antiserum with wide epitope reactivity, rabbits were immunized with a mixture of native and denatured hsp70 purified from bovine muscle by ATP-affinity chromatography. Screening for antibody specificity was performed by a "sandwich" enzyme linked immunosorbent assay (ELISA). Immunoprecipitation and immunoblotting analyses demonstrated that the polyclonal antiserum obtained by us and a monoclonal antibody raised against a different preparation of antigen recognized the same determinant on the native hsp70 molecule (inducible form). With a different specificity the polyclonal antiserum recognized only the denatured monomers of the other members of the hsp70 family. These results are discussed in relation to the immunological features of the hsp70 molecule and to the development of an immunoassay for the detection of hsp70 in cell and tissue extracts.
主要应激蛋白的hsp70家族由几个具有相似结构和功能特性的不同成员组成。为了获得具有广泛表位反应性的抗血清，用通过ATP亲和层析从牛肌肉中纯化的天然和变性hsp70混合物对兔子进行免疫。通过“夹心”酶联免疫吸附测定（ELISA）进行抗体特异性筛选。免疫沉淀和免疫印迹分析表明，我们获得的多克隆抗血清和针对不同抗原制剂产生的单克隆抗体识别天然hsp70分子（诱导型）上的相同决定簇。多克隆抗血清以不同的特异性仅识别hsp70家族其他成员的变性单体。结合hsp70分子的免疫学特征以及用于检测细胞和组织提取物中hsp70的免疫测定方法的开发对这些结果进行了讨论。
