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°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 .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 20.5. 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.

At automated fluorescence polarization immunoassay for the determination of vancomycin levels in serum was evaluated. The vancomycin assay is a homQgeneous -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 b'ioassay 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 ,ug 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.

A high-pressure liquid chromatographic method for the determination of norfloxacin or ciprofloxacin concentratioits in body fluids was developed and compared with a standard bioassay. The high-pressure liquid chromatographic assay utilizes a reverse-phase Cls 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. Cortelation coefficients with the bioassay were 0.966 for norfloxacin and 0.952 for ciprofloxacin.Norfloxacin and ciprofloxacin are new fluoroquinolones with appreciable antimicrobial activity against a broad spectrum of both gram-positive and gram-negative organisms (1,2,5,7,10,13,14). In a recent review article, Hooper and Wolfson (8) reported on the pharmacology and toxicity of the fluoroquinolones in humans. They reported that in patients with severe renal failure, the half-life of norfloxacin is prolonged, and for both ciprofloxacin and norfloxacin there is evidence that clearance involves both glomerular ifitration and renal tubular secretion. Because the accurate measurement of concentrations in body fluids would allow the adjustment of dosage regimens to avoid toxicity in patients with renal failure and to obtain therapeutic levels in patients with decreased gastrointestinal absorption, we developed a high-pressure liquid chromatography (HPLC) assay for norfloxacin and ciprofloxacin and compared it with a standard agar well diffusion assay.The HPLC was performed with a Waters solvent delivery system (model 6000A; Waters Associates, Inc., Milford, Mass.) at a rate of 2 ml/min and a Waters fluorescence detector (model 420) with excitation and emission wavelengths of 278 and 456 nm, respectively. Analysis was performed on a reverse-phase ,uBondapak C18 column (10 ,um; 15 cin by 3.9 mm; Waters). Samples were injected with a Rheodyne syringe loading sample injector (model 7125) fitted with a 20-j.l loop (Alltech Associates, Inc., Applied Science Div., State College, Pa.). The mobile phase for HPLC analysis was an acetonitrile-tetrabutyl ammonium hydroxide-phosphate buffer (pH 3.0) which was prepared by adding 1.67 ml of o-phosphoric acid (85%; J. T. Baker Chemical Co., Phillipsburg, N.J.), 15 ml of tetrabutylammonium hydroxide (1.54 M; Sigma Chemical Co., St. Louis, Mo.), and 100 ml of acetonitrile to 1 liter of deionized water. The mobile phase was filtered and degassed before use and was stable for at least 2 days when stored at 4°C. All deionized water used in the preparation of reagents was further purified by passage through a Nanopure water purification system (Sybron/Barrstead, Boston, Mass.).Serum and urine samples used in this study were obtained from leukemia or bone marrow transplant patients receiving norfloxacin or ciprofloxacin as well as other antibiotics for which various concentration determinations in serum had been ordered as part of their clinical management. When possible, sera were analyzed by both methods on the same * Corr...

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 difussion 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.Ticarcillin (oa-carboxy-3-thienylmethylpenicillin) is a semisynthetic penicillin exhibiting a high degree of activity against Pseudomonas aleruginosa and other gram-negative organisms (5,9,10,12,15,18). In addition to being used to treat proven Pseudomonas infections, ticarcillin is also used empirically in the immunocompromised host. Often, in both cases, ticarcillin is concurrently administered with aminoglycosides or cephalosporins or both. Toxicity associated with the use of penicillins is usually minimal; however, central nervous system side effects can be seen with high levels in serum (14). Although not required routinely, ticarcillin levels in serum should be monitored in patients with renal insufficiency, particularly when other beta-lactam antibiotics are being coadministered, and during the treatment of infections caused by organisms with high MICs to the antibiotic.Traditionally, ticarcillin levels in serum have been determined by microbiological assays, primarily the agar well diffussion (AWD) bioassay (1,5,6,8,11,13,(15)(16)(17)(18). Although cost-effective, these assays often lack the specificity and precision associated with biochemical assays or immunoassays for determining levels of antibiotics and require a minimum of 8 h of incubation, eliminating the possibility of dosage adjustment within the dosage interval. High-pressure liquid chromatographic (HPLC) procedures for determining ticarcillin concentrations have been reported for the quantitation of ticarcillin and other selected penicillins and cephalosporins in pharmaceutical preparations (4) and for the measurement of ticarcillin in serum and urine (7). While both assays are useful for the quantitation of ticarcillin, the former does not employ an internal standard to reference for quantitation, and the latter assay is cumbersome for application in a clinical laboratory. We therefore developed an HPLC method with a reverse-phase C18 pLBondapack column with internal standard quantitation which is applicable for routine use in the clinical laboratory. MATERIALS AND METHODSReagents. Ticarcilli...