Abstract
Type 1 diabetes (T1D) results from the destruction of pancreatic insulin-producing beta cells and is strongly associated with the presence of islet autoantibodies. Autoantibodies to tyrosine phosphatase-like protein IA-2 (IA-2As) are considered to be highly predictive markers of T1D. We developed a novel lateral flow immunoassay (LFIA) based on a bridging format for the rapid detection of IA-2As in human serum samples. In this assay, one site of the IA-2As is bound to HA-tagged-IA-2, which is subsequently captured on the anti-HA-Tag antibody-coated test line on the strip. The other site of the IA-2As is bound to biotinylated IA-2, allowing the complex to be visualized using colloidal gold nanoparticle-conjugated streptavidin. For this study, 35 serum samples from T1D patients and 44 control sera from non-diabetic individuals were analyzed with our novel assay and the results were correlated with two IA-2A ELISAs. Among the 35 serum samples from T1D patients, the IA-2A LFIA, the in-house IA-2A ELISA and the commercial IA-2A ELISA identified as positive 21, 29 and 30 IA-2A-positive sera, respectively. The major advantages of the IA-2A LFIA are its rapidity and simplicity.
Highlights
Type 1 diabetes (T1D) is an autoimmune disease characterized by the destruction of pancreatic beta cells within the islets of Langerhans
We describe the development of a double antigen bridging lateral flow immunoassay (LFIA) for the detection of IA-2 autoantibodies (IA-2As) in human serum samples
Assay optimization Since our IA-2A LFIA was designed as a double antigen bridging assay, two tags were required to implement this format with a control serum produce a purple band in the control area on the device, but no band in the test area
Summary
Type 1 diabetes (T1D) is an autoimmune disease characterized by the destruction of pancreatic beta cells within the islets of Langerhans. In the course of this autoimmune process, autoantibodies are generated against several beta-cell antigens, e.g. insulin, glutamic acid decarboxylase (GAD65), tyrosine phosphatase-like protein (IA-2) and zinc transporter 8 (ZnT8). Other methods for the detection of IA-2As use enzyme-linked immunosorbent assays (ELISAs) and time-resolved fluorescence assays, in which the immobilized antigen captures autoantibodies from the sample and detection is achieved using labeled antigen [12,13,14]. Even though these assays do not require radiolabeled compounds, commercially available ELISAs are relatively timeconsuming and expensive and still need specialized equipment. There is a need for assays for detecting autoantibodies to IA-2 that are rapid, easy to use, inexpensive and implementable in most clinical laboratories without any special expertise or equipment
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