Mass spectrometry analysis of bovine tuberculins - 3R potential in batch potency testing.

Tuberculin purified protein derivatives (tuberculin PPDs) are heat-treated, protein-enriched products of lysed mycobacteria. Tuberculin PPDs reveal a delayed hypersensitivity in individuals earlier sensitized to mycobacteria and are thereby used to detect tuberculosis. For batch potency testing of tuberculins, the European Pharmacopoeia advises intradermal injection of products into previously sensitized guinea pigs. Batch potency is reflected by the size of the resulting skin lesions as compared to a reference tuberculin. This procedure is quite compromising and results are highly variable, often requiring test repeats. In a proof of concept study, we evaluated the suitability of a combination of liquid chromatography and mass spectrometry (LC-MSE) to record qualitative protein profiles of bovine PPD tuberculin concentrates. Six batches of bovine PPD tuberculin concentrates from three manufacturers and the WHO International Standard for PPD of Mycobacterium bovis tuberculin were studied. In total, 35 proteins were identified by LC-MSE followed by MS database search. Eight proteins were consistently found in all batches of all bovine tuberculin PPD products. Our results suggest that LC-MSE can be used for the analysis of PPD tuberculin concentrates and that it can be further developed in the future into quantitative MS methods for batch analysis, i.e. consistency testing.