Development of An Automated High-Throughput Dried Blood Spot Assay to Facilitate Large Scale Screening for Type 1 Diabetes Risk

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Type 1 diabetes (T1D) is a chronic autoimmune disease that affects millions of children and adults in the US. Up to 40% of T1D patients are diagnosed in the emergency room with life-threatening diabetic ketoacidosis (DKA), resulting in severe clinical complications and a substantial economic burden. Programs that screen for blood-borne markers of early-stage T1D (anti-islet cell autoantibodies) can significantly diminish the incidence of DKA and improve patient quality of life. However, large-scale implementation of such screening campaigns is hindered by the high cost and low throughput of radioimmunoassay (RIA), the gold-standard method for measuring anti-islet autoantibodies. Moreover, a significant portion of screening program costs are associated with phlebotomy, cold-chain storage and shipping. To address this, Enable Biosciences aims to develop a cost-effective, multiplex, high- throughput and non-radioactive immunoassay to diagnose T1D and screen for individuals at risk for T1D using low-cost easy-to-collect dried blood spots (DBS). The successful development of this high- throughput dried blood spot T1D assay can substantially increase the effectiveness of general population- based T1D screening programs. In a JDRF- and Stanford-supported initiative, we were gratified that a serum-based Enable assay achieved the highest performance for anti-GAD, anti-IA2 and a close second-to-highest performance for anti-insulin antibodies in the recent blinded 2018 Islet-cell Autoantibody Standardization Program (IASP) competition out of 50 participating laboratories. The IASP data were generated with an automated Enable serum assay using a custom-made Enable analyzer created in collaboration with Hamilton Robotics. Furthermore, we showed that our manual DBS- based Enable assay correlated strongly with a serum-based assay (R=0.90-0.96). The concordance as evaluated by Cohen’s test showed kappa between 0.85-0.89. The positive and negative agreements for all three aforementioned autoantibodies range from 85%-99%. Notably, we also demonstrated less than 10% degradation of assay signals when DBS cards were stored at 37°C for 4 weeks. Building on these solid technical achievements, which we consider as our “Phase I-like Results” described in detail in this proposal, we aim to further automate our DBS assay procedures for high-throughput functionality including DBS elution and DBS eluent testing using diverse samples, such as those from Stanford University School of Medicine. We expect to deliver an integrated DBS T1D autoantibody assay (elution/testing automation device and reagent kits) to serve large screening centers and CLIA- laboratories. We further plan to obtain CE Mark classification and FDA approval to further decentralize DBS T1D autoantibody testing to be able to address the important needs of communities, clinicians and researchers worldwide.