Shichida Y, Taniguchi Y, Kuwata O, Fukada Y, Yoshizawa T, Horiuchi S, Takeichi M
Department of Biophysics, Faculty of Science, Kyoto University, Japan.
Exp Eye Res. 1989 Feb;48(2):281-93. doi: 10.1016/s0014-4835(89)80077-6.
The protein moiety of chicken iodopsin, R-photopsin, was purified from the chicken retina using a sucrose flotation method followed by two steps of column chromatography. Apparent molecular weights of R-photopsin and scotopsin (the protein moiety of chicken rhodopsin), which was partly purified in the process of purification of R-photopsin, were estimated to be 34,000 and 36,000, respectively, by sodium docecylsulfate-polyacrylamide gel electrophoresis. Using the purified R-photopsin as an antigen, four kinds of hybridoma cells which secreted monoclonal antibodies specific for R-photopsin and iodopsin were prepared. The antibodies thus obtained reacted with neither other chicken cone visual pigments nor rhodopsin as analyzed by immunoblots and immunoprecipitation methods. All the monoclonal antibodies stained the majority of the cone outer segments in chicken retina, while an antiserum raised against cattle rhodopsin stained the rod outer segments as well as some cone outer segments in the retina.
鸡视锥细胞视蛋白（R-视蛋白）的蛋白质部分通过蔗糖浮选法从鸡视网膜中纯化出来，随后经过两步柱色谱法。通过十二烷基硫酸钠-聚丙烯酰胺凝胶电泳估计，R-视蛋白和暗视蛋白（鸡视紫红质的蛋白质部分，在R-视蛋白纯化过程中部分纯化）的表观分子量分别为34,000和36,000。以纯化的R-视蛋白为抗原，制备了四种分泌对R-视蛋白和视锥细胞视蛋白具有特异性的单克隆抗体的杂交瘤细胞。通过免疫印迹和免疫沉淀法分析，所获得的抗体与其他鸡视锥细胞视觉色素或视紫红质均无反应。所有单克隆抗体均能对鸡视网膜中的大多数视锥细胞外段进行染色，而针对牛视紫红质产生的抗血清则能对视网膜中的视杆细胞外段以及一些视锥细胞外段进行染色。
