QuickFISH Background

QuickFISH is a rapid, simple slide based assay that provides a definitive pathogen identification, directly from a positive blood culture, in just 20 minutes.
It is ideally suited to providing accurate diagnosis of bacteraemia, candidaemia and sepsis in the clinical microbiology lab.

QuickFISH compares favourably with biochemical tests, amplification tests and MALDI-TOF. Considering commercially available molecular tests, QuickFISH has the fastest turn around time.” 1.

The QuickFISH system is a novel molecular diagnostic system based on the patented PNA-FISH technology. The principle of the assay is a simple one, enhancing the well-known technique of Fluorescent In Situ Hybridisation (FISH) by incorporating the use of Peptide Nucleic Acids (PNA).

Peptide Nucleic Acid (PNA) molecules are DNA mimics in which the negatively charged sugar phosphate backbone found in DNA and RNA is replaced with a non-charged polyamide or “peptide” backbone, while retaining the nucleotide bases that form the basis of the genetic code.

This means that a PNA molecule will hybridise according to the same base-pairing rules as any other nucleic acid. However as the PNA molecule is unaffected by electrostatic repulsion between negatively charged backbones, it will bind more rapidly and specifically than a comparable DNA or RNA molecule. This means that probes composed of PNA can be shorter, and their hydrophobic backbone allows them to cross cell membrane with ease.

Revolutionary PNA Probe Technology

Probe-quencher complexes enable fast and highly specific targeting of pathgen rRNA via florescence In Situ Hybridisation (FISH).

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Room Temp: Target specific PNA Probe-Quencher complexes quench florescence signal.

Hybridisation at 55°: When heated to 55°C, probes and quenchers separate and probes hybridise to target rRNA, if present.

Cooling to Room Temperature: During cooling, probe-quencher complexes reform. Probes hybridised to target rRNA and NOT quenched enabling florescence visualisation of target pathogen.

 

The PNA-FISH technology uses these characteristics to allow pathogens to be identified quickly and accurately. By using PNA-FISH probes with a complementary sequence to the pathogen’s ribosomal RNA, a specific and sensitive reaction enables whole cell visualisation of the target pathogen(s) using fluorescence microscopy. The assay requires minimal sample preparation as cells don’t need to be lysed to isolate genetic material and generates visual results that match Gram-stain morphology. Furthermore the QuickFISH platform has eliminated the need for wash steps. As a definitive pathogen identification can be obtained in 20 minutes, using a simple protocol, the QuickFISH technique is ideal for use in any clinical microbiology laboratory.

Quick and Easy QuickFISH Procedure

3 Easy Steps, 5 minutes hands on time, 20 minutes turn-around.

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Step 1: Fix – Fix 10µL of blood culture sample to QuickFISH slide.

Step 2: Hybridise – Add PNA reagents and hybridise for 15 minutes at 55°C.

Step 3: Examine – View results on florescence microscope (60x or 100x Oil Objective).

1.    Deck, M.K. et alRapid detection of Enterococcus spp. Direct from blood cultures using Enterococcus QuickFISH method: a multicentre investigation – in press in Diagnostic Microbiology and Infectious Disease. Diagn Microbiol Infect Dis 2013, Article in Press DOI: 10.1016/j.diagmicrobio.2013.12.004