PCR Mycoplasma Detection Kit

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Catalogue No: abx298017
Price: US$507.50
(Size: 100 rxns)
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PCR Mycoplasma Detection Kit

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Datasheet
PCR Mycoplasma Detection Kit is designed to detect the presence of mycoplasma contamination by PCR in biological samples, such as cell culture supernatants, which can be tested directly without DNA extraction. Highly specific primers have been designed to amplify a fragment of 16S rRNA coding DNA that is conserved across all commonly-known mycoplasma species. This kit is able to detect as low as 20 copies of the mycoplasma genome and only detects mycoplasma DNA, not eukaryotic or bacterial DNA. The kit includes an optimized supermix and primer, ultrapure water and positive control template. The kit provides a very easy to use, simple, fast (within 2 hours), specific and sensitive PCR-based mycoplasma detection method.

Kit components:
  • 2X PCR Myco SuperMix: 1 ml
  • Myco Primer Mix: 40 µl
  • Myco Positive Control Template: 40 µl
  • Myco-Free Water: 1 ml


Target PCR Mycoplasma Detection Kit
Storage Store at -20 °C for up to two years.
Availability Shipped within 10-20 working days.
Note This product is for research use only.
Directions for use Sample Preparation:

  • Adherent cells: Cells should be cultured for 2-3 days with ~80% confluence. Transfer 40 µl of cell culture supernatant to a PCR tube and incubate at 95 °C for 10 minutes in a thermocyler.
  • Suspension cells: Cells should be cultured for 2-3 days with a density of ~106 cells/ml. Transfer the cell culture to a microcentrifuge tube and centrifuge at 2000 × g for 1 minute. Transfer 40 µl of cell culture supernatant to a PCR tube and incubate at 95 °C for 10 minutes in a thermocyler.
  • Serum: Dilute serum 20-fold. Transfer 40 µl of diluted serum to a PCR tube and incubate at 95 °C for 10 minutes in a thermocyler.

Samples can be stored at -20 °C for 1 month before or after heat treatment. Cells that have been cultured for more than 4 days should be diluted 10-fold before being tested in PCR.

PCR Setup – Reaction Components:

Component Volume
Template 2 µl
Myco Primer Mix 0.4 µl
2X Myco PCR SuperMix 10 µl
Myco-Free Water7.6 µl
Total Volume 20 µl

The provided Myco-Free Water can be used as a negative control and Myco Positive Control Template as the positive control.

PCR Amplification – Thermal Cycling Conditions:

Number of Cycles Temperature Time per Cycle
1 cycle 94 °C 4 min
35 cycles 94 °C 30 seconds
60 °C 30 seconds
72 °C30 seconds
1 cycle 72 °C5 min

Results and Analysis:
  1. Agarose gel electrophoresis: Load 10 µl of PCR product onto a 1.5% agarose gel and resolve by electrophoresis.
  2. Results Interpretation: Compare with the positive and negative control. The size of the positive band is about 350 bp.
Notes:

  • To avoid mycoplasma contamination, the PCR reactions should be carried out in a designated PCR area and all reagents should be kept on ice during set-up. Face masks are recommended, as the oral cavity contains mycoplasma which may contaminate samples.
  • Before use, completely thaw all components and mix well.
  • Antibiotics such as penicillin, streptomycin, or serum in cell culture samples will not interfere with the assay.
  • This product can be used together with abx098886 Mycoplasma Prevention Reagent and/or abx098884 and abx098885 Mycoplasma Elimination Reagents.
Research Articles on PCR Mycoplasma Detection Kit


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