EVALUATION OF MICROBIAL INHIBITOR TESTS WITH INDICATOR IN MICROTITRE PLATES BY PHOTOMETRIC MEASUREMENTS

Routine testing with the widespread use of microbial inhibitor tests w ith indicator in microtitre plates is generally evaluated visually in 2-3 steps (negative, questionable, positive). The objectives in develo ping a photometric evaluation were: To have the possibility of an obje ctive evaluation in order to increase the repeatability of test result s both within and between laboratories, and to fulfill the necessity o f an evaluation method which allows more subtle differentiation of res ults in validation studies and during the development/modificiation of inhibitor tests. Four negative and 4 positive (4 mu g penicillin/kg) controls were tested on each test plate. As control/reference samples, 1 batch of lyophilized milk was used for the whole examination period in order to allow a comparison of data of all test plates. After the incubation each well of the plates was read by an ELISA Reader (550 nm /690 nm measurement/reference wavelength). In order to obtain a standa rdized scale, the difference between the mean absorption values of neg ative and positive controls was set as 100 % and the absorption values of the wells of the corresponding test plates calculated in 10 % step s, with respect to the control values (''relativated absorption in %'' ). Due to the frequency distribution of relativated absorption values within visually evaluated steps, samples classified either as negative or positive are separated whereas some overlapping within the visuall y evaluated step questionable exists. Examples of the application of t he method of relativated absorption within a validation programme (rep eatability between and shelf life of test kit batches), and during met hodological development (choice of suitable beta-lactamase preparation s and activities for the identification of beta-lactam antibiotics) ar e given.