一种新的考马斯亮兰R—250固相染料结合法测蛋白质含量的方法

A new solid-phase dye-binding method for proteindetermination with coomassie breiliant Blue R-250

本文介绍了一种用考马斯亮兰Ｒ—２５０因相染料结合法测蛋白质含量的方法。将蛋白样品点在滤纸上，甲醇固定，考马斯亮兰Ｒ—２５０染色，脱色，用洗脱液洗脱下与染料结合的蛋白质，在波长６１０ｎｍ处测其光密度（ｏｐｔｉｃａｌｄｅｎｓｉｔｙ．Ｏ．Ｄ）值。方法简便，所用样品少，灵敏度高，检测线性范围在０．１ｇ／Ｌ～５ｇ／Ｌ间，重复性较好，批内变异３．５６％～６．６７％，批间变异１．４８％～７．０２％。样品稳定，染色后的滤纸可保存一个月，洗脱液体放置４８ｈ颜色不变。受干扰物质少是本法的一个特点，仅受ＳＤＳ、强碱等少数几种物质的干扰，不受叠氮钠、硫酸铵、ＴｒｔｏｎＸ－１００等物质的干扰。

This is a protein determination method which involves the binding of CoomassieBrilliant Blue R - 250 to immobilized protein. Put Protein solution to filter paper and fix it withabsolute methamol. Coomassie Brilliant Blue R - 250 dyes the protein. Rinse the filter paper,wash the dye binding proteins in filter paper and measure the optical density (O. D)of elutedprotein-dye complexes. The method is simple and convenient. The analysis requires small sam-ple volume and has high sensitivity. The linear range of assay are between 0. 1g/L and 5g/L.The assay has good reproducibility. The coefficients of variation in the group are 3. 56%￣6.67%and between groups are 1. 48%￣7. 02 %. The method is stable. The dyed filter papers canbe stored for 1 month without sensible changes in the O. D. values. The eluted solution hasgood color stability for 48 hr. Only a few reagents such as sodium dodecyl sulfate.alkali havesome interference effections on this detecting method. However,hardly 10 reagents,like sodiumazide,ammonium sulfate. Tritonx - 100 etc. have not interference effections.