Objective
Mass screening for islet autoantibodies (IAbs) in the general population is being conducted globally to prevent type 1 diabetes (T1D). A highly efficient and easy to conduct IAb screening assay across laboratories is needed. A most common laboratory method, enzyme-linked immunosorbent assay (ELISA) for detecting a major islet autoantibody (IAb), insulin autoantibodies (IAA), is still not available although many laboratories made a great effort for the last three decades but failed due to technique difficulties. IAA is most critical for early prediction of risk of T1D development, especially in young children since IAA is the earliest IAb in majority with a very high rate of positivity that has been demonstrated in multiple T1D preventive clinical trial studies. IAA assay is most difficult, compared with three other IAb assays, and has the widest variation of sensitivity and specificity between the participant laboratories in multiple diabetes autoantibody workshop led by NIH and Immunology of Diabetes Society. All ELISA methods for IAA, so far, don’t work at all. Our laboratory, serving as the NIH/NIDDK reference autoantibody laboratory, has recently developed a novel bridging ELISA to measure IAA with excellent sensitivity and specificity. In this application, we are proposing to optimize and validate this new novel ELISA-IAA assay with a large cohort of samples from new onset T1D patients, pre-T1D subjects, and age/gender matched healthy controls, compared with a current standard method of radio-binding assay (RBA) for its sensitivity, specificity, and disease prediction. In addition, we will set up this new ELISA assays, with the same assay format, for three other IAbs including GADA, IA-2A, and ZnT8A, which are warranted upon the excellent results from the IAA ELISA assay. Furthermore, we are going to combine all 4-IAb ELISA together into one to build up a multiplex ELISA assay, which will be able to screen a complete panel of all 4 T1D IAbs, fit for large scale population screening.
Background Rationale
Appearance of islet autoantibodies (IAbs) in the peripheral blood is currently the most reliable biomarker for T1D, both presymptomatic T1D and progression to clinical disease. The incidence of T1D is rapidly increasing and double every 20 years, especially in young children. In the US alone, 1.6 million people have T1D and as many have IAbs or preclinical T1D. Insulin autoantibodies (IAA) are most critical for early prediction of risk of T1D development, especially in young children since IAA is the earliest IAb in majority with a very high rate of positivity, which has been well-documented in multiple prospective new-born cohort studies including Diabetes Autoimmunity Study in the Young (DAISY) and The Environmental Determinants of Diabetes in the Young (TEDDY) study. IAA assay is most difficult, compared with three other IAb assays (GADA, IA-2A, and ZnT8A) and has the widest variation of sensitivity and specificity between the participant laboratories in Islet Autoantibody Standardization Program (IASP) workshop. For the last three decades, many laboratories have made a great effort trying to find a way with the most common laboratory method, enzyme-linked immunosorbent assay (ELISA) for measuring IAA, but technical challenges have hindered success. All ELISA methods for IAA, so far, don’t work at all. Our laboratory serves as the NIH/NIDDK reference autoantibody laboratory for most major initiatives in T1D research, e.g., TrialNet, TEDDY, ITN, T1DGC. We have recently developed a novel bridging ELISA to measure IAA with excellent sensitivity and specificity, compared with a current standard method of radio-binding assay (RBA). The designing of bridging ELISA was based on our high-affinity IAA assay with electrochemiluminescence (ECL) technology which discriminates high-affinity (disease predictive) from low-affinity (non-disease predictive) IAA. We expect the bridging ELISA-IAA, like the ECL-IAA, will be more predictive than the RBA-IAA in population-based screening. Mass screening for IAbs in the general population is being conducted globally to prevent T1D. A highly efficient and easy to conduct IAb screening assay across laboratories is needed. In this proposal, we will validate this new bridging ELISA-IAA, adopt this bridging ELISA for three other IAbs (GADA, IA-2A, and ZnT8A), and further build a multiplex ELISA to combine a complete panel of all 4 IAbs into one, which will be expected to be the best tool for the general population screening with high sensitivity/specificity, high throughput, easy to use and low cost.
Description of Project
Type 1 diabetes (T1D), the immune-mediated form of diabetes, is one of the most common and serious long-term diseases in children. Over 1.6 million people in the U.S. alone are affected by T1D and the incidence doubles every 20 years, especially in young children. Millions of people may benefit from screening for yet undiagnosed or presymptomatic islet autoimmunity and from early treatment. Screening of the general population children for pre-symptomatic T1D has being conducted globally to prevent T1D. A highly efficient and easy to conduct pancreatic islet autoantibody (IAb) screening assay across laboratories is needed. A most common laboratory method, enzyme-linked immunosorbent assay (ELISA) for detecting a major IAb, insulin autoantibodies (IAA), is still not available although many laboratories made a great effort but failed due to technique difficulties. IAA is most critical for early prediction of risk of T1D development, especially in young children since IAA is the earliest IAb in majority with a very high rate of positivity that has been demonstrated in multiple clinical trial studies. Our laboratory, serving as the NIH/NIDDK reference autoantibody laboratory, has recently developed a novel ELISA method to measure IAA with excellent sensitivity and specificity, compared with a current standard method of radio-binding assay (RBA). In this application, we are proposing to optimize and validate this new novel ELISA-IAA assay with a large cohort of samples from new onset T1D patients, pre-T1D subjects, and age/gender matched healthy controls. We expect this simple IAA assay will offer the simplest way, as a commonly used method of traditional ELISA existed in all laboratories, with high sensitivity and specificity, high efficiency, easy to use, and lower cost than all existing assays including current standard method of RBA. It will facilitate all clinical and research laboratories for T1D risk screening and clinical diagnosis. With the same assay format, we will set up the same ELISA assays for three other IAbs including GADA, IA-2A, and ZnT8A, which are warranted upon the excellent results from the IAA ELISA assay. Furthermore, we are going to combine all 4-IAb ELISA together into one to build up a multiplex ELISA for this proposal, which will be able to screen a complete panel of all 4 T1D IAbs in one step, fit for large scale population screening with high sensitivity/specificity, high throughput, easy to use, and low cost.
Anticipated Outcome
Mass screening for islet autoantibodies (IAbs) in the general population is being conducted globally to prevent type 1 diabetes (T1D). A highly efficient and easy to conduct IAb screening assay across laboratories is needed. Insulin autoantibodies (IAA) are most critical for early prediction of risk of T1D development, especially in young children since IAA is the earliest IAb in majority with a very high rate of positivity that has been demonstrated in multiple prospective T1D clinical trial studies. IAA assay is most difficult, compared with three other IAb assays, and has the widest variation of sensitivity and specificity between the participant laboratories in Islet Autoantibody Standardization Program (IASP) workshop. A most common laboratory method, enzyme-linked immunosorbent assay (ELISA) for detecting a major IAb, insulin autoantibodies (IAA), is still not available although many laboratories made a great effort for last three decades but failed due to technique difficulties. We have recently developed a novel bridging ELISA to measure IAA with excellent sensitivity and specificity where the validation with a large-scale cohort is warranted. We expect this simple ELISA IAA assay will offer the easiest way, as a commonly used method existing in all laboratories. It will facilitate all clinical and research laboratories for T1D risk screening and clinical diagnosis. With the same assay format, we will set up the same ELISA assays for three other IAbs including GADA, IA-2A, and ZnT8A, and further build a simple multiplex ELISA combining all 4 IAbs into one fit for the high-throughput population screening, which we expect to achieve a simplest multiplex assay for a complete panel of IAbs with high sensitivity and specificity, high efficiency, easy to use, and low cost for universal screening of the general population or high-risk groups for T1D risk with the goal of early diagnosis, treatment, and eventually prevention of diabetes onset.
Relevance to T1D
Type 1 diabetes (T1D), the immune-mediated form of diabetes, is one of the most common and serious long-term diseases in children. Over 1.6 million people in the U.S. alone are affected by T1D and the incidence doubles every 20 years, especially in young children. Millions of people may benefit from screening for yet undiagnosed or presymptomatic islet autoimmunity and from early treatment. Appearance of islet autoantibodies (IAbs) in the peripheral blood is currently the most reliable biomarker for T1D, both presymptomatic T1D and progression to clinical disease. With a more efficient and economical assay format, this simple ELISA assay technology will greatly benefit all clinical and research laboratories for T1D risk screening and clinical diagnosis, and facilitate the general population screening for T1D risk and greatly enhance diabetes prevention efforts.