การพัฒนาระบบอิเล็กโทรโฟริซิส เพื่อวิเคราะห์โพลีมอร์ฟิคเอนไซม์บางชนิด สำหรับงานนิติเวชศาสตร์ในประเทศไทย / สมพร โชติเกษมสุข= Development of electrophoretic system for typing of some polymorphic enzymes for forensic science in Thailand
Electrophoresis is a biochemical method applicable for polymorphic enzyme analysis for blood individualization. The object of this work is to evaluate the most appropriated electrophoretic system for typing human blood phosphoglucomutase (PGM), erythrocyte acid phosphatase (EAP) and esterase D (EsD). The evaluation was performed on agarose and polyacrylamide gel electrophoretic systems and polyacrylamide gel isoelectric focusing. The latter was by vertical midget electrophoresis (VM – IEF), horizontal mini IEF (HM – IEF) and horizontal chamber electrophoresis (H – IEF). PGM1 isozymes could be resolved by agarose-starch gel electrophoresis at pH 7.41 into 3 subtyes : PGM1 1, PGM1 2 and PGM1 2-1. Polyarylamide gel electrophoresis (PAGF) with continuous system at pH 7.4 or with discontinuous system at pH 8.9 failed to resolve the isoenzyme. The best resolution with 4 isozymes of 10 subtypes : 1+, 1-, 1+1, 2+, 2-, 2+2-, 1+2-, 1-2+ and 1-2- was obtained from isoelectric focusing (IEF) on polyacrylamide gel at pH 5-7 by H – IEF and at pH 5.0 – 6.5 with 1.2% EPPS by HM – IEF. EAP isozymes could be resolved into 3 subtypes : A, B and BA by isoelectric focusing on polyacrylamide gel at pH 4-8 by H – IEF and at pH 5.0 – 6.5 by HM – IEF. By the same system with 3.14% HEPES addition, EAP and EsD could be typed simultaneously. The latter could be resolved into 5 subtypes : EsD 1, EsD 2, EsD 2.1, EsD 5.1 and EsD 5.2. In stability study, PGM was the most stable polymorphic enzyme. The ages of these three enzymes which were still typeable were 105, 21 and 12 days, respectively. Gene frequency of PGM, EAP and EsD obtained by isoelectric focusing on polyacrylamide gel1+ at pH 5-7 and 4-8, determined from 250 samples of Thai population were PGM11+ (0.598), PGM11- (0.126), PGM12+ (0.162), PGM12- (0.116), PA (0.292), PB (0.708), EsD1 (0.688) and EsD5 (0.006) respectively. Discriminating power of PGM1, EAP and EsD were 0.79, 0.57 and 0.59, respectively. The total discriminating power resulted from these three polymorphic enzyme typing was 0.96.