哺乳动物细胞裂解物中新霉素磷酸转移酶活性的定量原位凝胶电泳测定法。

Quantitative in situ gel electrophoretic assay for neomycin phosphotransferase activity in mammalian cell lysates.

作者信息

Fregien N, Davidson N

出版信息

Anal Biochem. 1985 Jul;148(1):101-4. doi: 10.1016/0003-2697(85)90633-5.

Abstract

A method for the assay of neomycin phosphotransferase activity in eucaryotic cell lysates is described. Total cytoplasmic proteins are fractionated in nondenaturing polyacrylamide gels and then allowed to react in situ with [gamma-32P]ATP and kanamycin. The reaction products are detected by blotting to phosphocellulose paper and autoradiography. The assay is linear with protein concentration and sensitive enough to detect expression in transient assays.

摘要

本文描述了一种用于检测真核细胞裂解物中新霉素磷酸转移酶活性的方法。将总细胞质蛋白在非变性聚丙烯酰胺凝胶中进行分级分离，然后使其与[γ-32P]ATP和卡那霉素在原位反应。通过将反应产物印迹到磷酸纤维素纸上并进行放射自显影来检测。该检测方法与蛋白质浓度呈线性关系，灵敏度足以在瞬时检测中检测到表达。