Basu A, Modak M J
J Biol Chem. 1987 Feb 15;262(5):2369-73.
We have carried out photoaffinity labeling of the ras p21 protein, a ras oncogene product, with [alpha-32P]GTP. Based on our studies, a sensitive, rapid, and specific assay for the detection of multiple forms of ras p21 has been developed. The specificity of this protocol is shown by (a) sensitivity of affinity labeling of ras p21 to known inhibitors of GTP binding and (b) immunoprecipitation of affinity labeled protein with anti-ras p21 serum. Detection and semiquantitation of ras p21 by this method is accomplished in less than 24 h and requires as little as 100,000 cells or about 5 mg of tissue sample from skin tumor, liver, and mammary tumor tissues. Furthermore, using this approach, we were able to detect the selective loss of one species of ras p21 in transplanted Morris hepatoma cells.
我们用[α-32P]GTP对ras p21蛋白（一种ras癌基因产物）进行了光亲和标记。基于我们的研究，已开发出一种灵敏、快速且特异的检测多种形式ras p21的方法。该方法的特异性体现在：（a）ras p21的亲和标记对已知GTP结合抑制剂敏感；（b）用抗ras p21血清对亲和标记蛋白进行免疫沉淀。用这种方法检测和半定量ras p21在不到24小时内即可完成，所需细胞量低至100,000个，或来自皮肤肿瘤、肝脏和乳腺肿瘤组织的组织样本约5毫克。此外，使用这种方法，我们能够检测移植的莫里斯肝癌细胞中一种ras p21的选择性缺失。
