Specific Aspects of Tuberculin Products Standardisation

Abstract

According to the existing international practice, reference materials, which are used to assess specific activity of purified tuberculin products, are meant to be used for several decades and are therefore characterised by high stability. For instance, PPD-S tuberculin produced in the USA in 1944 has been used ever since as the international reference standard of purified tuberculin and is stored as lyophilisate in ampoules (PPDT) containing 5000 IU each. The Russian PPD-L-2 industry reference standard is made from material produced by the Leningrad Research Institute of Vaccines and Sera in 1973. It has been used for many years to control production batches of PPD-L purified tuberculin because of its high stability that is regularly confirmed in large-scale studies using various experimental models in accordance with the requirements of the World Health Organisation. The aim of the study was to evaluate the long-term stability of the substance of industry reference standard of purified tuberculin (PPD-L-2 IRS) by determining its specific activity relative to the international standard of purified tuberculin, and determine the feasibility of using this substance for the preparation of lyophilised reference product samples. Materials and methods: PPD-L-2 IRS specific activity was determined relative to the PPDT international standard in accordance with the procedure specified in the monograph FS.3.3.1.0023.15 of the State Pharmacopoeia of Russian Federation (14 edition) using guinea pigs vaccinated with various BCG substrains or sensitized by virulent mycobacteria («live» or «inactivated») in accordance with the recommendations of the WHO (WHO TRS 45, 1987). Results: the analysis of the obtained data showed that there were 3–4-fold differences in PPD-L-2 IRS relative potency depending on the BCG substrain used for guinea pigs vaccination (animal sensitization model). This effect of the titration model manifested itself when comparing specific activity of tuberculins obtained by various methods of tuberculoprotein precipitation (PPD-L-2 and PPDT), i.e. different in antigen composition. The specific activity of the previously established dose of PPD-L-2 IRS was shown to be equivalent to the international reference standard in animals sensitized with mycobacteria tuberculosis. Conclusions: the results of PPD-L-2 IRS specific activity assessment obtained in this study are consistent with the data obtained during development and certification of this reference material in the 1980s and confirm the long-term stability of the intermediate powder product of the Russian reference standard of purified tuberculin. At the same time, the production of a freeze-dried form of the IRS, in addition to being economically feasible, would rule out some potential errors that are inevitable during annual preparation and control of the reference standard dilutions, and would make it possible to spare the substance which would improve prospects for the future long-term use of PPD-L as an industry reference standard.