This Funding Opportunity Announcement (FOA) encourages applications from institutions/organizations proposing the development and/or validation of targeted mass spectrometric assays (e.g. Multiple Reaction Monitoring) for proteins and peptides of primary interest to the type 1 diabetes research community [e.g. glucagon and other pro-glucagon derived peptides, C-peptide, insulin, pro-insulin, Glycated CD59, Islet Amyloid Polypeptide (IAPP), Chromogranin A (CgA), and chromogranin B (CgB)]. The proposed assays should be highly reproducible, easily transferable to other laboratories, and validated in human plasma or serum. This might also require the development of appropriate community standards, and reference materials when not already available.
Many assays in basic and clinical science research rely exclusively on antibodies. However, there are no widely accepted guidelines or standardized methods to determine the validity of these reagents. Furthermore, many recent publications have highlighted the limitations of commercial antibodies including failure to detect the intended target. The rigor and reproducibility of many assays commonly used by researchers in the field of type 1 diabetes could be substantially improved applying Mass Spectrometry (MS) instead of relying only on antibodies.
For some analytes, mass spectrometric assays that are more specific and less variable than previous measurements have been already developed. For example, for insulin and c-peptide the feasibility of using Liquid Chromatography-Mass Spectrometry (LC-MS) in a high-throughput clinical laboratory was demonstrated. However, these methods might need further validation. For glucagon and other proglucagon derived peptides, a highly reproducible assay that combines the use of a monoclonal antibody for enrichment and mass spectrometry for the detection and quantitation was developed. However, the monoclonal antibody used in the enrichment step was developed in house and is not easily available to the scientific community. More recently, a mass spectrometers manufacturer used a combination of solid-phase extraction (SPE) and µElution to develop an assay for glucagon in human plasma. Although this assay might need further validation, it demonstrated that it is possible to reach a Limit of Detection in the low pg/mL with very simple and inexpensive fractionation steps prior to a targeted mass spectrometry experiment like a Multiple Reaction Monitoring (MRM) experiment. Novel interfaces, that can be placed in the front-end of the mass spectrometer, can also be used for reducing chemical noise and matrix interference, leading to targeted MS assays that are more robust. Furthermore, these types of assays with simple pre-fractionation steps are in many cases easy to multiplex.
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