Related Experiment Videos
Microplate phosphocellulose binding assay for aminoglycoside-modifying enzymes.
Antimicrobial Agents and Chemotherapy
|December 1, 1986
Summary
This study optimized the phosphocellulose binding assay for detecting aminoglycoside-modifying enzymes (AMEs) using microdilution plates. The new method is faster, more convenient, and cost-effective for screening numerous bacterial isolates.
Area of Science:
- Biochemistry
- Microbiology
- Enzymology
Background:
- Aminoglycoside-modifying enzymes (AMEs) are crucial for bacterial resistance.
- Accurate and efficient detection of AMEs is vital for clinical and research microbiology.
Purpose of the Study:
- To adapt and validate a microplate-based phosphocellulose binding assay for AME detection.
- To improve the speed, convenience, and cost-effectiveness of screening for AMEs.
Main Methods:
- Modified the phosphocellulose binding assay using microdilution plates and multichannel micropipettes.
- Prepared and stored aminoglycoside substrates in microdilution plates for screening and enzyme subclassification.
- Compared the microplate assay with the conventional tube reaction assay using 32 bacterial isolates from nine species.
Main Results:
- The microplate assay demonstrated comparable radioactive counts to the conventional assay.
- Equally correct identifications of AMEs were achieved in all tested bacterial isolates.
- Modifications enabled rapid, convenient, and less expensive examination of large organism collections.
Conclusions:
- The microplate-based assay is a validated and effective method for detecting and subclassifying aminoglycoside-modifying enzymes.
- This optimized assay facilitates high-throughput screening of bacterial populations for AME presence.
- The enhanced assay offers significant advantages in terms of speed, cost, and convenience for microbiological laboratories.