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Common Reference-Based Tandem Mass Tag Multiplexing for the Relative Quantification of Peptides: Design and Application to Degradome Analysis of Diphtheria Toxoid.

ABSTRACT: Currently, animal tests are being used to confirm the potency and lack of toxicity of toxoid vaccines. In a consistency approach, animal tests could be replaced if production consistency (compared to known good products) can be proven in a panel of in vitro assays. By mimicking the in vivo antigen processing in a simplified in vitro approach, it may be possible to distinguish aberrant products from good products. To demonstrate this, heat-exposed diphtheria toxoid was subjected to partial digestion by cathepsin S (an endoprotease involved in antigen processing), and the peptide formation/degradation kinetics were mapped for various heated toxoids. To overcome the limitations associated with the very large number of samples, we used common reference-based tandem mass tag (TMT) labeling. Instead of using one label per condition with direct comparison between the set of labels, we compared multiple labeled samples to a common reference (a pooled sample containing an aliquot of each condition). In this method, the number of samples is not limited by the number of unique TMT labels. This TMT multiplexing strategy allows for a 15-fold reduction of analysis time while retaining the reliability advantage of TMT labeling over label-free quantification. The formation of the most important peptides could be followed over time and compared among several conditions. The changes in enzymatic degradation kinetics of diphtheria toxoid revealed several suitable candidate peptides for use in a quality control assay that can distinguish structurally aberrant diphtheria toxoid from compliant toxoids.

SUBMITTER: Michiels TJM

PROVIDER: S-EPMC8176455 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:ObjectiveTetanus toxoid, reduced diphtheria toxoid, and acellular pertusiss (Tdap) vaccine is recommended during each pregnancy, regardless of prior receipt. Data on reactogenicity and immunogenicity, particularly after repeated Tdap, are limited. We compared local injection-site and systemic reactions and serologic response following Tdap in (1) pregnant and nonpregnant women and (2) pregnant women by self-reported prior Tdap receipt.Study designPregnant women (gestational age 20-34 weeks) and nonpregnant women receiving Tdap were enrolled in this observational study. Injection-site and systemic reactions were assessed for one week post-vaccination. Pertussis toxin, filamentous hemagglutinin, pertactin, fimbriae, tetanus and diphtheria specific IgG antibody titers were determined by standardized enzyme-linked immunosorbent assay at baseline and 28 days post-vaccination. Reactogenicity and serologic responses were compared by pregnancy status, and within pregnant women by self-reported prior Tdap receipt.Results374 pregnant and 225 nonpregnant women were vaccinated. Severe local or systemic reactions or "any" fever were uncommon (≤3% for both groups). Moderate/severe injection-site pain was significantly higher in pregnant (17.9%) versus nonpregnant (11.1%) women, but did not prompt a healthcare visit. Proportions of other moderate/severe or any severe reactions were not significantly higher in pregnant compared to nonpregnant women. Moderate/severe (including pain) and severe reactions were not significantly higher in pregnant women receiving repeat versus first-time Tdap. Antibody titers increased from baseline to post-vaccination for all vaccine antigens in pregnant and nonpregnant women; post-vaccination titers against pertussis toxin and filamentous hemagglutinin were significantly higher in nonpregnant versus pregnant women (p < 0.01).ConclusionTdap was well-tolerated in pregnant and nonpregnant women. Pregnant women were more likely to report moderate/severe pain at the Tdap injection-site compared with nonpregnant women, but did not necessitate medical visits. Prior Tdap receipt did not increase occurrence of moderate/severe local or systemic reactions in pregnant women. Serologic responses to all vaccine antigens were robust. Clinical Trial Registration@ClinicalTrials.gov. NCT02209623. https://clinicaltrials.gov/ct2/show/NCT02209623.

Dataset Information

Common Reference-Based Tandem Mass Tag Multiplexing for the Relative Quantification of Peptides: Design and Application to Degradome Analysis of Diphtheria Toxoid.

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