A Label-free Electrochemiluminescent Immunosensor For Glutamate Decarboxylase Antibody Detection On AuNPs Supporting Interface

Diabetes mellitus is a chronic endocrine disease with the characters of hyperglycemia and glucose metabolism disorder.The incidence of type 1 diabetes mellitus（T1DM）,a genetically immune-mediated form of diabetes which was caused by dysfunction ofβ-cell for insulin secretion.Autoimmune diabetes of adult late onset（LADA）is also caused by islet beta cell function damage of immune-mediated disease,the two types of diabetes often lead to an absolute lack of insulin and have a high incidence in our country.The number of patients has doubled in last 20 years.Therefore,timely screening,reasonable monitoring and prevention of complications are of great significance.Some autoimmune markers including glutamic acid decarboxylase antibody（GADA）and islet cell antibody（ICA）and insulin autoantibodies（IAA）,protein tyrosine phosphatase antibody（anti-IA2）,and zinc transporter 8 autoantibody（ZnT8A）etc can be found in serum of those persons whom are in risk of autoimmune injury for their pancreatic islet cells.The abnormal rise of GADA is most closely related with T1 DM,and can be employed to distinguish LADA from Type II diabetes.Since the 1990 s,the glutamate decarboxylase has been confirmed universally as a vital target antigen in pancreaticβ-cell immune injury,thus,the GADA has attracted intensive attention in recent years.Its assay is of great value for early diagnosis,monitoring and prevention of T1 DM and LADA.At present,ELISA method mainly applied to the detection of GADA,but it is costly and time-consuming and so on.Therefore,it is of great significance to develop a simple and fast method for the detection of GADA in diabetic patients.In this paper,we described a novel lable-free electrochemiluminescent（ECL）immunosensor for the detection of glutamate decarboxylase antibody（GADA）in which Au nanoparticles（AuNPs）had pre-functionalized the indium tin oxide（ITO）glass to support the sensing interface.The preparation of the basal electrode only need a simple two-step drop coating,a thin polymer of hydrolyzed 3-aminopropyl-trimethoxysilane,and the AuNPs gel on the ITO substrate.The AuNPs not only enhanced the ECL signal of luminol,but also acted to immobilize the GAD65 to build the sensing host,then to block non-specific binding sites with BSA,the immunosensor is acquired.The performance of GAD65 immunosensor was characterized by scanning electron microscopy（SEM）,electrochemical impedance spectroscopy（EIS）and so on.On optimal conditions,this immunosensor can test GADA by using luminol as ECL probe,the result showed that the decreased ECL intensity has a good linear regression toward the logarithm of GADA concentration in the range of 0.30 ng·mL^-1to 50 ng·mL^-1with a detection limit of 0.10 ng·mL^-1（1.5pM）.In addition,this immunosensor possessed great specificity,high sensitivity and low cost,suitable for high-throughput immunoassay.