Development of a novel multiplex electrochemiluminescent-based immunoassay to aid enterotoxigenic Escherichia coli vaccine development and evaluations

Abstract

Enterotoxigenic Escherichia coli (ETEC) is a leading cause of bacterial diarrhea both among children in low and middle income countries and in travelers to these regions. Although there are several approaches to develop an effective vaccine for ETEC, no licensed vaccines are currently available. The most advanced ETEC vaccine candidates include multiple colonization factors along with the heat labile toxin B subunit. In the absence of known correlates of protection, and to understand the mechanism of protection, monitoring immune responses to a majority of the vaccine associated antigens using various types of samples is needed. Unfortunately, a traditional ELISA is time consuming, labor intensive and requires substantial amounts of antigens and sample volumes. To address these constraints, we developed and validated a novel high throughput electrochemiluminescent (ECL) - based multiplex immunoassay using Meso Scale Discovery (MSD) platform for analyzing immune responses to ETEC antigens. The ETEC multiplex ECL assay is an 8-plex assay which includes the ETEC colonization factor antigens (CFA/I, CS1, CS2, CS3, CS5 and CS6) along with the two subunits of heat labile toxin (LTA and LTB). Our data suggested that a single dilution of sample provides a quantifiable result for a wide range of sample titers. To compare ETEC multiplex ECL with ELISA, we carried out assays using the same antigens with the two immunoassay platforms using a common sample set of serum and ALS (antibodies in lymphocyte supernatant) specimens. The MSD platform achieved excellent correlations with ELISA for the antigens tested, consistently detecting comparable antibody levels in the samples. The ETEC multiplex ECL can serve as a fundamental platform in evaluating performances of candidate ETEC vaccines in future field trials.

Systemic inflammatory markers in neuropathic pain, nerve injury, and recovery

“…U-PLEX plate custom biomarker multiplex assay kits (Meso Scale Diagnostics LLC, USA) were custom designed to detect 18 selected cytokines/chemokines related to the gene expression data with high sensitivity ( Supplementary Table 1 ) [ 12 , 16 ]. Processing followed the standard manufacturer protocols.…”

Linking inherent O-Linked Protein Glycosylation of YghJ to Increased Antigen Potential

“…SoftMax Pro 7 software was used to fit measured intensity values (arbitrary units) as a function of serum concentration. Endpoint titers at 0.4 units above background were read from the fitted curve as described in Chakraborty et al, 2019(Chakraborty et al, 2019.…”

Multiplexed Immunosensors and Immunoarrays

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