研究目的
To evaluate a new molecular application of peptide nucleic acid–fluorescence in situ hybridization (PNA-FISH) with Staphylococcus-specific molecular PNA probes for the rapid detection of common pathogens causing endophthalmitis.
研究成果
The study demonstrates a proof of concept for using PNA-FISH with Staphylococcus-specific probes to rapidly identify pathogens in endophthalmitis isolates, with high accuracy and a detection time of 20 minutes, suggesting potential clinical benefits for improving diagnosis and treatment outcomes.
研究不足
Small sample size, case scenarios with polymicrobial infections, use of artificial medium instead of intraocular fluid, potential low sensitivity with minimal bacterial load, and the need for modification for intraocular fluid samples.
1：Experimental Design and Method Selection:
An experimental proof-of-concept study was conducted at the microbiology laboratory using the PNA-FISH technique with QuickFISH protocol for hybridization.
2：Sample Selection and Data Sources:
Stored culture-positive staphylococci endophthalmitis isolates from prior vitreous samples (n=15) and negative controls (broth, n=5) were used, selected based on prior identification via conventional microbiology tests and stored frozen.
3：List of Experimental Equipment and Materials:
Includes fluorescence microscope, incubator/slidestation, pipettes, QuickFISH slides, fixation solutions (QuickFIX-1 and QuickFIX-2), PNA probes (yellow and blue), blood culture bottles, thioglycollate broth, and skimmed milk for storage.
4：Experimental Procedures and Operational Workflow:
Inoculum was prepared to 1x10^5 CFU/mL, samples were fixed and hybridized on slides at 55°C for 15 minutes, and results were read qualitatively by two independent observers using a fluorescence microscope at 360x magnification.
5：Data Analysis Methods:
Qualitative assessment by observers for detection based on shape and color, with no statistical analysis mentioned.
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