Identification of clinical isolates of gram-negative nonfermentative bacteria by an automated cellular fatty acid identification system.

An automated cellular fatty acid (CFA) bacterial identification system, Microbial Identification System (MIS; Microbial ID, Newark, Del.), was compared with a conventional system for the identification of 573 strains of gram-negative nonfermentative bacteria. MIS identifications were based exclusively on the CFA composition following 22 to 26 h of growth at 28 degrees C on Trypticase soy agar. MIS identifications were listed with a confidence measurement (similarity index [SI]) on a scale of 0 to 1.0. A value of greater than or equal to 0.5 was considered a good match. The MIS correctly listed as the first choice 478 of 532 (90%) strains contained in the data base. However, only 314 (59%) had SI values of greater than or equal to 0.5. Of the 54 strains in which there was not agreement, 37 belonged to the genera Acinetobacter, Moraxella, or Alcaligenes or were Pseudomonas pickettii. Reproducibility studies suggest that SI variation is most likely a function of a difference in culture age at the time of analysis, which is due to the relatively low temperature and time of incubation. Other discrepancies were attributable to insufficiently characterized library entries or an inability to differentiate chemotaxonomically closely related species. The MIS, as the first automated CFA identification system, is an accurate, efficient, and relatively rapid method for the identification of gram-negative nonfermentative bacteria. The development of a CFA library with the media and incubation conditions routinely used for the isolation of clinical pathogens could further decrease the identification time and provide an increase in accuracy.

Determination of norfloxacin and ciprofloxacin concentrations in serum and urine by high-pressure liquid chromatography.

A high-pressure liquid chromatographic method for the determination of norfloxacin or ciprofloxacin concentrations in body fluids was developed and compared with a standard bioassay. The high-pressure liquid chromatographic assay utilizes a reverse-phase C18 column, an internal standard, and fluorescence detection, with reproducibility studies yielding coefficients of variation ranging from 0.6 to 3.7% and 0.9 to 2.7% for norfloxacin and ciprofloxacin, respectively. Correlation coefficients with the bioassay were 0.966 for norfloxacin and 0.952 for ciprofloxacin.

Determination of ticarcillin levels in serum by high-pressure liquid chromatography.

A high-pressure liquid chromatographic method for determining the concentrations of ticarcillin in serum was developed and compared, with 93 patient sera, to a standard agar well diffusion bioassay. For analysis, serum plus temocillin, the internal standard, were extracted with chloroform-n-amyl alcohol and back extracted into phosphate buffer. A reverse-phase C18 column and an ammonium acetate-methanol mobile phase were used with detection at 242 nm. Reproducibility studies yielded coefficients of variation ranging from 2.4 to 4.7% for low, mid, and high controls. Although cefoxitin, cephalothin, and cefuroxime exhibited retention similar to that of ticarcillin, a wide range of commonly administered antibiotics and other drugs did not interfere. The high-pressure liquid chromatographic assay is an accurate, reproducible method for determining the concentration of ticarcillin in serum during multiple antibiotic therapy or when rapid results are required.

Evaluation of an automated fluorescence polarization immunoassay for vancomycin.

An automated fluorescence polarization immunoassay for the determination of vancomycin levels in serum was evaluated. The vancomycin assay is a homogeneous competitive inhibition immunoassay based on changes in fluorescence polarization that occur with antibody binding. This assay was compared with a liquid chromatographic assay and an agar well diffusion bioassay method by using clinical serum specimens and controls. Linear regression analysis of the data obtained on clinical specimens by the three methods resulted in correlation coefficients of 0.97 for the fluorescence polarization immunoassay versus the liquid chromatographic assay (n = 60), 0.90 for the fluorescence polarization immunoassay versus the bioassay (n = 57), and 0.90 for the liquid chromatographic assay versus the bioassay (n = 57). Repetitive analysis of control sera containing 7, 35, and 75 micrograms of vancomycin per ml by the fluorescence polarization immunoassay yielded coefficients of variation of 3.0, 1.7, and 2.3, respectively. No interference was demonstrated in spiked hemolytic, lipemic, or icteric sera, and the assay was free of matrix effects. The automated fluorescence polarization immunoassay system offers a rapid, efficient, and accurate method for monitoring vancomycin serum levels for both toxicity and efficacy.