DETECTION OF AUTOANTIBODIES GAD 65

AULANNI’AM ¹,  FATCHIYAH ¹,  DJOKO W. S. ²   SUTIMAN  B. ¹

¹Faculty of  Mathematic and Natural Sciences , Brawijaya University and

 ²⁾  Medical Faculty , Brawijaya University

  

ABSTRACT

            Since the importance of anti-GAD65 assays to predict IDDM (Insulin Dependent Diabetes Mellitus), therefore we observed the possibility of getting the alternative reagents and method for faster, more accurate and low cost diagnostic. In this observe, normal rabbits induced by anti-GAD65 from rat’s antisera, to produce antibodies to anti-GAD65 (anti-GAD65 antibodies). Antibodies collected and purified by 50% SAS, then we conjugated the purified antibodies with Alkaline Phosphatase (AP). The result of dot blotting which use GAD enzyme from bovine brain as antigen and rabbit’s antibodies are able to recognize anti-GAD65 both from rat’s antisera and pre-diabetic patient’s sera. This evidence explain that there is a possibility to use GAD65 enzyme and anti-GAD65 antibodies conjugated AP as diagnostic reagent for IDDM prediction. GAD65 enzyme as diagnostic reagent has optimum sensitivity in 1/50 (enzyme : phosphate buffer) and anti-GAD antibodies conjugated AP has optimum sensitivity in 1/150 (antisera : TBS) as secondary antibodies.

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