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Mycoplasma Detection


Mycoplasma infections are relatively common in laboratory cell cultures; it has been estimated that between 5% and 35% of all cell cultures are infected. Mycoplasmas have been shown to alter the growth rate of cells in culture, induce chromosomal aberrations, influence amino acid and nucleic acid metabolism and cause membrane aberrations. Several methods have been developed to detect mycoplasma including direct culture in special growth media, enzyme-linked immunoassay, immunofluorescence staining, PCR, biochemical detection and fluorescent nucleic acid stains.

ABP Biosciences has developed three methods for fast and sensitive detection of Mycoplasma infection: MycoCheck™ Mycoplasma PCR Detection Kit, MycoCheck™ Mycoplasma Luminescent Detection Kit and MycoCheck™ Mycoplasma Stain Kit.


MycoCheck™ Mycoplasma PCR Detection Kit

The MycoCheck™ Mycoplasma PCR Detection Kit allows for fast and reliable identification of mycoplasma contamination in cell cultures. Mycoplasma DNA in the cell culture supernatant is amplified via PCR and visualized using gel electrophoresis. In addition to the short detection process (less than 2 hours), the easy handling and high sensitivity makes this Mycoplasma PCR Detection Kit a convenient tool for routine examination of cell cultures and media. The kit contains an optimized PCR master mix and a positive control. The primers in the PCR master mix are highly specific to the conserved rRNA region in the mycoplasma genomes and can detect all well-known mycoplasma genera, including the commonly encountered ones in cell cultures, such as M. orale, M. hyorhinis, M. laidlawii, M. salivariumM. arginini, M. fermentans, M. hominis, and M. pneumoniae. Mycoplasma positive samples can be easily recognized by a distinctive PCR product ranging in size from 400 to 600 bp.

Figure 1. The PCR fragments of the commonly encountered mycoplasmas in cell cultures


MycoCheck™ Mycoplasma Luminescent Detection Kit

The MycoCheck™ Mycoplasma Luminescent Detection Kit allows for quick and reliable screening of cell cultures for contamination with mycoplasmas. The assay utilizes the activity of mycoplasma-specific enzymes and detects through the luminescence reaction catalyzed by ATP-dependent luciferase. The entire detection process has only two steps and only takes about 15 minutes. The first step is to add mycoplasma detection reagent A to the sample and perform the test after 5 minutes. The reading value is A. The second step is to add mycoplasma detection reagent B and perform the test after 10 minutes. The reading value is B. If the sample is contaminated by mycoplasma, its unique enzyme can convert the ADP into ATP. By measuring the level of ATP in a sample both before and after the addition of the mycoplasma detection reagent B, a ratio can be obtained which is indicative of the presence or absence of mycoplasma.

Figure 2. The kinetics of ATP generation in the presence of M.hyorhinis contamination


MycoCheck™ Mycoplasma Stain Kit

The MycoCheck™ Mycoplasma Stain Kit provides an ultrasensitive, rapid and simple fluorescence microscopic assay for the visual identification of mycoplasma infection in laboratory cell cultures. In order to detect mycoplasma, the fluorescent MycoFluor reagent is added directly to the culture medium, with or without cells, and the stained sample is then examined under a fluorescence microscope. The excitation and emission spectra of MycoFluor reagent bound to dsDNA are similar to dsDNA bound to DAPI, with excitation maxima around 350-360nm and emission maxima around 450-460nm. MycoFluor reagent can be excited either with a xenon mercury arc lamp or a UV laser and is detected through a blue filter.

Mycoplasma staining with MycoFluor reagent appears as a fine particulate or filamentous staining over the cytoplasm at 100X magnification. Nuclei of the cells are also brightly stained by this method and thereby act as endogenous positive control for the staining procedure.

Figure 3. The Jurkat cell culture stained with MycoCheck™ Mycoplasma Stain Kit

To order

Buy Cat.No. Product Name Unit Price
A058-1 MycoCheck™ Mycoplasma PCR Detection Kit 20T $110
A058-2 40T $150
A059-1 MycoCheck™ Mycoplasma Stain Kit 2 mL $110
A059-2 10 mL $165
A066-1 MycoCheck™ Mycoplasma Luminescent Detection Kit 20T $150
A066-2 100T $450