Viability PCR, also named v-PCR or vPCR, is an evolution of

PCR PCR or pcr may refer to: Science * Phosphocreatine, a phosphorylated creatine molecule * Principal component regression, a statistical technique Medicine * Polymerase chain reaction ** COVID-19 testing, often performed using the polymerase chain r ...

. Through the use of a simple pre-treatment of the sample by the means of specific intercalating photo-reactive reagents it's possible to neutralize the DNA of dead cells. As a result, only DNA from live cells will be detected by PCR. This approach expands a lot the analytical scope of PCR procedures. The capability to detect only living cells become very important, because in key applications is more important to know the amount of live cells, than the total cell level. Examples of this are: food and water quality control, infectious diseases diagnostic, veterinary applications, ecological dynamics... The first referenced work about this analytical approach was in 2003, Norwegian researchers suggest the use of Ethidium Monoazide, an azide form of

Ethidium Bromide Ethidium bromide (or homidium bromide, chloride salt homidium chloride) is an intercalating agent commonly used as a fluorescent tag ( nucleic acid stain) in molecular biology laboratories for techniques such as agarose gel electrophoresis. I ...

, which was used in other analytical fields as

Flow Cytometry Flow cytometry (FC) is a technique used to detect and measure physical and chemical characteristics of a population of cells or particles. In this process, a sample containing cells or particles is suspended in a fluid and injected into the fl ...

as a candidate for viability PCR. However, the main important advances were done by Nocker and colleagues, which demonstrated in successive works the potential of this technology and also suggested

Propidium monoazide Propidium monoazide (PMA) is a photoreactive DNA-binding dye that preferentially binds to dsDNA. It is used to detect viable microorganisms by qPCR. Visible light (high power halogen lamps or specific LED devices) induces a photoreaction of the ...

as a better reagent for vPCR. This field still is in development, from 2003 up to 2015, the scientific evidences about the applicability of vPCR are stacking, nowadays main efforts are focused in procedure optimization. Since a simple reagent mix with the sample, photo-activation and subsequent PCR not always shows expected results, each procedure needs some optimization. Up to now the main improvements has been : - Improving the efficiency of photo activation: early procedures were based on high power halogen lamps which overheated the samples and don't ensured constant light dose, these home made solutions have been replaced by led based instrument

https://www.qiagen.com/us/shop/sample-technologies/dna/dna-preparation/blu-v-system/] - The use of long PCR amplicons as targets. - The increase of temperature during dark incubation. Through combining different optimizations strategies and controlling the analytical bias, nowadays the vPCR becomes a powerful analytical tool.

References

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