BLU-V Viability PMA Kit
For differentiation between live and dead microorganisms
QIAGEN PMA Reagent uses the DNA-intercalating agent Propidium Monoazide (PMA) to permit differentiation of live and dead microorganisms at DNA level.
PMA selectively enters dead cells and upon photo-activation intercalates and binds covalently to DNA and strongly inhibits its amplification in subsequent PCR reactions. The resulting signal from dead cell DNA then significantly shifts to a higher CT value, thereby providing a clear distinction between live and dead cells.
The BLU-V Viability PMA Kit is intended for molecular biology applications in food, animal feed, and pharmaceutical product testing. These products are not intended for the diagnosis, prevention, or treatment of a disease.
PMA-based viability PCR clearly differentiates live and dead cell DNA. PMA selectively enters dead cells and upon photo-activation, intercalates and binds covalently to DNA, strongly inhibiting its amplification in subsequent PCR reactions. The resulting signal from dead cell DNA then significantly shifts to a higher CT value, thereby providing a clear distinction between live and dead cells (see figure PMA inhibits amplification of dead cell DNA).
Propidium Monoazide (PMA) is a photoactivatable, chemical compound, that has found wide acceptance in molecular biology applications as a technology for live/dead cell differentiation in microbiological organisms.
Following photoactivation with a defined wavelength, PMA intercalates and binds covalently to DNA. Subsequent amplification of the modified DNA is inhibited, resulting in a reduction in the amplification signal.
PMA is unable to pass through intact biological membranes. Subsequently, DNA from living microorganisms, which have intact membranes, is protected from PMA modifications and is therefore detectable by PCR. Membranes of dead microorganisms lose their protective functionality, and PMA can enter these cells and modify their DNA. Modified DNA of dead microorganisms is inhibited from detection by PCR, resulting in a reduced amplification signal.
Bacterial species are prepared and treated with PMA. Following activation of the compound by illumination with a specific wavelength in the BLU-V System, the reagent is irreversibly bound to DNA of dead cells, masking it so that PCR amplification is prevented. Efficient suppression of amplification of such modified DNA allows preferential detection of live cells.
Live/dead differentiation can play an important role in procedures such as — hygiene testing to measure the success of decontamination processes, water testing, to distinguish between live and dead Legionella for regulatory compliance, and to monitor the efficiency of pathogen killing.
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