Development of amilase and acid phosphatase detection kit

Amylase is an enzyme which is abundant in the saliva and it is used as saliva biomarker. The acid phosphatase is abundant semen and it is used to detect of semen. Saliva and semen detections are particularly evidence of the sexual assault cases in forensic medicine. In this thesis, semen (acid phosphatase) and saliva (amylase), which are biological evidences of sexual assault cases, diagnostic kit has developed with a new detection method. The principle of method for acid phosphatase detection is changing colour with reaction of diazonium salt and naphtol which occurs after hydrolyzation of alpha naphthyl phosphate with enzyme. Starch iodine test was used for the detection of amylase. Reaction solutions as prepared for both tests were absorbed to special gel matrix as support medium. For the detection sensitivity, 10.000 to 100.000 dilutions were performed in semen samples, 100 to 1000 dilutions were performed in saliva samples. For time-based activity assay, 10 µl saliva and semen was dropped to 5mm2 cotton fabrics and allowed to dry. Samples were dissolved in distilled water and analyzed at 0, 1, 6, 12, 24, 36, 48, 60, 72th hours. Acid phosphatase detection in semen and amylase detection in saliva samples can be performed up to 1/70.000 and 1/900 dilution, respectively. In time-based activity assay, activity was detected even in 72th hour which is reported in literature that enzyme lost majority of activity. Matrix-assisted method that developed in this study is more practical and sensitive compared to the classical method.