Study On Preparation And Application As A Reference Material Of Gamma-glutamyltransferase

The method of metrological traceability of values for catalytic concentration of enzymes is now widely recognized as the combination of reference materials, reference measurement procedures and network of reference laboratories. Reference materials that are value-assigned using an internationally agreed upon protocol play an important role in this system and now reference materials or calibrators have come to our attention in our country. Although quality control and quality assessment are developed in clinical laboratories, reference materials are not widely used at present. The first reason is that all calibrators are imported and very expensive. The second reason is that some questions on the calibrator commutability are present in different measurement systems. So the propose of this study is to achieve calibrators made in our country.In this study, Gamma-glutamyltransferase(GGT, EC2.3.2.2) was purified from pig kidney. We get the GGT sample solution with high specific activity up to 124 U/mg and high purity because of the single band on electrophoresis gel through homogenization, solubilization with sodium deoxycholate, precipitation ammonium sulfate and incubation with papain, and further fine purify with chromatography on sephadex G-25 to desalt, deae sepharose and superdex 75. The molecule of sample protein is 21 kD, and a hydrophilic subunit of previous protein.We studied the stability and commutability of the enzyme in different matrix after having set up the reference measurement procedure. With the elementary observation, at 4℃and -20℃, the enzyme keep its activity concentration stable in CRM matrix and self-made matrix in 120 days, but at 37℃, it loss 5% activity concentration after 120 days. Comparing the sample in human serum matrix, which loss about 15.2% at 4℃and 39% at 37℃, the stability of sample in CRM matrix and self-made matrix is excellent. If we compare the two kinds of general methods to the reference method, only the sample in CRM matrix with high value go beyond the 95% confident interval. It show the good commutability of the sample among the the two kinds of general methods and the reference method, especially in CRM matrix and self-made matrix. It could be used as a reference material.