Bacterial Drug Sensitivity Identified by Immunoflow Cytometry

Antibiotic sensitivity or resistance of Staphylococcus aureus can be reliably differentiated by flow cytometry when labeled with nucleic acid dye.

An alternative approach to differentiate between methicillin-resistant S. aureus (MRSA) and methicillin-susceptible S. aureus (MSSA) can be achieved by labeling with a S. aureus-specific antistaphylococcal protein A antibody instead of nucleic acid dyes.

Scientists at the Cleveland Clinic, (OH ,USA) analyzed a total of 103 clinical isolates of S. aureus including 49 MRSA and 54 MSSA, obtained from the Cleveland Clinic clinical microbiology laboratory. These were isolates from blood, respiratory, tissue, and wound cultures, and were all obtained from different patients. Labeled bacterial cells were analyzed in a Micro PRO flow cytometer using a sampling volume of 0.25 mL.

All 103 isolates were correctly identified as S. aureus by producing a signal when labeled with the anti-S. aureus protein A antibody. Ten coagulase-negative staphylococci and two enterococci used as controls did not produce a signal with the antibody. A batch of 12 samples could be analyzed in 6.5 hours, which includes 4 hours of incubation in the presence of oxacillin, and 1 hour of this being hands-on time. The Micro PRO flow cytometer (Advanced Analytical Technologies, Inc.; Ames, IA, USA) used in the assay allows for random access testing, whereby individual samples can be analyzed independently. Although sample preparation would be more efficiently done in batches, the actual flow cytometry itself does not require batch testing, and a clinical laboratory has the option of using this technology in real-time.

The cost per sample of this immunoflow cytometry assay is estimated to be about USD 11 per sample, compared with about USD 24 by flow cytometry using a peptide nucleic acid (PNA) probe, and USD 45 for the same identification using a commercially available real-time polymerase chain reaction (PCR) assay. The costs of commercially available simple flow cytometers capable of analyzing particles the size of bacteria are estimated to be between USD 60,000 and USD 100,000. The authors concluded that immunoflow cytometry is a viable method of rapidly identifying S. aureus and differentiating MRSA from MSSA, and comes with many of the advantages of flow cytometry and microorganism-specific labeling associated with using PNA probes. The study was published on December 27, 2011, in the European Journal of Clinical Microbiology and Infectious Diseases.

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