Automated system for analytical microbiology V: calibration lines for antibiotics

The accuracy of an automated system for the microbiological assay of antibiotics was increased by improvement attendant to connection to an on-line computer. The system was used to investigate the suitability of four forms of interpolation formulas by assaying for chlortetracycline and erythromycin. The calibration lines were prepared as point-to-point straight-line approximations and as cubic equations. Cubic equations through four calibration points were preferred. Since the automated system was a four-channel instrument, a separate response line was prepared for each channel. Combining the four response lines into one could substantially degrade the accuracy and precision of assays. A new general equation relating the response of the test organism to concentrations of active materials was used to account for factors in addition to the antibiotic upon the dose-response line. Some of these factors were: diluents, growth substances, relative proportions of mixed antibiotics, pH and buffer capacities of the sample solution and assay broth, salts, and organic compounds in samples and not in standard solutions. The equation was used to show under what conditions the dose-response lines of mixtures and single-component antibiotics could be the same. It could also account for the nonspecific nature of turbidimetric assays. The equation showed assay biases to be caused not by differences in composition of antibiotics in standards and samples but by differences in other substances affecting growth of the test organism. A new dose-response line applicable to assays using Klebsiella pneumoniae was described.