Hough-Monroe L, Milstone L M
Veterans Affairs Medical Center, West Haven, Connecticut 06516.
Anal Biochem. 1991 Nov 15;199(1):25-8. doi: 10.1016/0003-2697(91)90264-t.
Covalently cross-linked protein is the key feature of the cornified envelope, an important marker of squamous differentiation. Enumeration of cornified envelopes is widely used to assess keratinocyte differentiation, but is tedious and subjective. We report here a rapid, objective, sensitive, and quantitative assay that measures total cross-linked protein in keratinocytes. The method is based on the resistance of cross-linked protein to solubilization in boiling SDS-beta ME, separation of cross-linked from soluble protein by collection of cross-linked protein on sheets of regenerated cellulose, binding of Coomassie blue to protein, and quantitation by scanning laser densitometry. There is excellent correlation between the cross-linked protein measured by this assay and the number of cornified envelopes.
共价交联蛋白是角质化包膜的关键特征，是鳞状分化的重要标志物。角质化包膜计数被广泛用于评估角质形成细胞分化，但既繁琐又主观。我们在此报告一种快速、客观、灵敏且定量的检测方法，用于测量角质形成细胞中的总交联蛋白。该方法基于交联蛋白对在沸腾的SDS-β巯基乙醇中溶解的抗性，通过在再生纤维素片上收集交联蛋白将交联蛋白与可溶性蛋白分离，考马斯亮蓝与蛋白结合，并通过扫描激光密度测定法定量。通过该检测方法测得的交联蛋白与角质化包膜数量之间存在极好的相关性。
