Clinical Lab Handbook

Container/Tube: SST

Alternate Container: Red Top

Specimen Volume: 5 mL of serum

Collection Instructions: Allow specimen to clot for 30 minutes. Spin specimen in centrifuge.

Centrifuge specimens and remove the serum or plasma from the cellular material within 2 hours of collection.

Enzyme-Multiplied Immunoassay Technique (EMIT®)

The VITROS TOBRA Reagent is a dual chambered package containing ready-to-use liquid reagents that are used in a twostep reaction to quantitatively measure tobramycin. Test sample is added to Reagent 1 containing tobramycin labeled with glucose-6-phosphate dehydrogenase (G6P-DH), followed by Reagent 2 containing antibody reactive to tobramycin, glucose-6-phosate and adenine dinucleotide (NAD). The assay is based on competition between tobramycin in the sample and tobramycin labeled with G6P-DH for antibody binding sites. G6P-DH activity decreases upon binding to the antibody, so tobramycin concentration in the sample can be measured in terms of G6P-DH activity. G6P-DH converts NAD+ to NADH, resulting in an absorbance change that is measured spectrophotometrically at 340 nm. Endogenous serum G6P-DH does not participate in the reaction since NAD+ functions only with the bacterial (Leuconostoc mesenteroides) enzyme conjugate employed in the assay.

Tobramycin is used in the treatment of serious infections caused by susceptible strains of gram-negative microorganisms and particular gram-positive organisms that are resistant to less toxic antibiotics. Tobramycin is safe and effective only in a narrow range of concentrations for a given indication. Exposure to high tobramycin concentrations for a prolonged period may cause renal impairment or ototoxicity. Serum or plasma tobramycin measurements are used in the diagnosis and treatment of tobramycin overdose and in monitoring levels of tobramycin to ensure appropriate therapy.

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