Mass Spectrometry-Based Quantification of the Antigens in Aluminum Hydroxide-Adjuvanted Diphtheria-Tetanus-Acellular-Pertussis Combination Vaccines

Abstract

Vaccines undergo stringent batch-release testing, most often including in-vivo assays for potency. For combination vaccines, such as diphtheria-tetanus-pertussis (DTaP), chemical modification induced by formaldehyde inactivation, as well as adsorption to aluminum-based adjuvants, complicates antigen-specific in-vitro analysis. Here, a mass spectrometric method was developed that allows the identification and quantitation of DTaP antigens in a combination vaccine. Isotopically labeled, antigen-specific internal standard peptides were employed that permitted absolute quantitation of their antigen-derived peptide counterparts and, consequently, the individual antigens. We evaluated the applicability of the method on monovalent non-adjuvanted antigens, on final vaccine lots and on experimental vaccine batches, where certain antigens were omitted from the drug product. Apart from the applicability for final batch release, we demonstrated the suitability of the approach for in-process control monitoring. The peptide quantification method facilitates antigen-specific identification and quantification of combination vaccines in a single assay. This may contribute, as part of the consistency approach, to a reduction in the number of animal tests required for vaccine-batch release.

Aluminum Adjuvants—‘Back to the Future’

“…Regulatory requirements for vaccines routinely include tightly controlled manufacturing and batch release processes. Two recent papers represent a tentative approach to extend the application of Raman spectroscopy and LC–MS, respectively, for the identification and quantification of tetanus, diphtheria, and pertussis (DTaP) antigens in aluminum-adjuvanted vaccines [ 94 , 95 ]. Raman spectroscopy demonstrated the potential for the identification and differentiation of complex vaccine products [ 94 ].…”

“…An LC–MS-based method was developed for the quantification of antigens in a final DTaP vaccine using antigen-specific signature peptides and isotopically labeled standards [ 95 ]. Individual antigens in a multivalent AH-adjuvanted DTaP vaccine were identified and specifically quantified using LC–MS, with quantification consistent with the concentrations supplied by the manufacturer.…”