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MP0035

Sigma-Aldrich

LookOut® Mycoplasma PCR Detection Kit

LookOut® Mycoplasma PCR Detection Kit

Optimized for use with JumpStart™ Taq DNA Polymerase, D9307.

Synonym(s):

PCR-based mycoplasma screening

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About This Item

UNSPSC Code:
12352207
NACRES:
NA.75
Pricing and availability is not currently available.

quality

Not suitable for clinical diagnostic use. Will not detect clinically relevant species such as M. pneumoniae and U. urealyticum

Quality Level

100

packaging

pkg of 1 kit

technique(s)

PCR: suitable

application(s)

detection
microbiology

compatibility

Optimized for use with JumpStart™ Taq DNA Polymerase, D9307.

storage temp.

2-8°C

General description

The LookOut® Mycoplasma PCR Detection Kit utilizes the polymerase chain reaction (PCR), which is established as the method of choice for highest sensitivity in the detection of Mycoplasma, Acholeplasma, and Ureaplasma contamination in cell cultures and other cell culture derived biologicals. Kit has been successfully tested for the ability to amplify Mycoplasma fermentans genomic DNA in concentrations of 20,000 to 20 copies.
The reaction tubes included with the kit are pre-coated with appropriate dNTPs, primers, and loading dye. Total assay time is greatly reduced compared to general protocols that require individual loading of reaction tubes.

Application

Mycoplasma PCR Detection Kit Video Protocol
The LookOut® Mycoplasma PCR Detection Kit has been used for screening of mycoplasma infection.[1][2]

Compatibility

The kit has been optimized for use with Product No. D9307. Any DNA polymerase with a 3′-5′ exonuclease activity is NOT recommended. These include Tli, Pfu,and Pwo. Those DNA polymerases that may be used include Taq, Tfl, and Tth.

Legal Information

JumpStart is a trademark of Sigma-Aldrich Co. LLC
LookOut is a registered trademark of Merck KGaA, Darmstadt, Germany

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Product No.
Description
Pricing

Storage Class Code

10 - Combustible liquids

Flash Point(F)

Not applicable

Flash Point(C)

Not applicable

Choose from one of the most recent versions:

Certificates of Analysis (COA)

Lot/Batch Number
BCCL4114
BCCL4119
BCCL8332
BCCK5837
BCCJ8830

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Optimized PCR-based Detection of Mycoplasma

Mycoplasma contamination of cell cultures is a serious issue impacting cell model validity. PCR testing for mycoplasma is an inexpensive, sensitive, and specific method for detecting contamination.

1 Review This action will navigate to reviews.

Questions

1–10 of 16 Questions  

  1. What is the full comprehensive list of all Mycoplasma species that the LookOut® Mycoplasma PCR Detection Kit, not test for? I am aware of the two outlined species, M. pneumoniae and U. urealyticum, but I am not sure on how these two are clinically similar

    1 answer

    1. This kit is intended for use as a mycoplasma screening and detection tool for cell culture use. It is not suitable for in vitro diagnostics use. While M. pneumoniae and U. urealyticum have been tested and found to be non-reactive, a full list of clinically associated mycoplasma species has not been determined. Please see the image below to review those species that have been tested using this kit.

      Please use the following link to review the product Technical Bulletin:
      https://www.sigmaaldrich.com/deepweb/assets/sigmaaldrich/product/documents/213/434/mp0035bul.pdf

      Helpful?

  2. What is the difference between product no. MP0040A (LookOut® Mycoplasma qPCR Detection Kit) and MP0035?

    1 answer

    1. The main differences between these kits are as follows:

      For MP0035:
      •Standard PCR
      •Primers/dNTPs precoated on tubes
      •Includes tubes
      •Taq Polymerase not included
      •Requires 40 cycles at 30 seconds

      For MP0040A:
      •qPCR only
      •Primers/dNTPs require addition to the reaction
      •Does not include tubes
      •Taq Polymerase included in primer/dNTP mix
      •Requires 45 cycles at 30 seconds
      MP0035 is suitable for 24 reactions, while MP0040A is suitable for 25 reactions.

      Helpful?

  3. What is the required DNA concentration of the purified sample supposed to be to ensure compatibility with the LookOut® Mycoplasma PCR Detection Kit, MP0035?

    1 answer

    1. The detection of Mycoplasma sp. with Product MP0035 - LookOut® Mycoplasma PCR Detection Kit requires less than 10 mycoplasma genomes per microliter of the sample, which is approximately 10 fg of DNA. It is important to ensure that the sample does not contain more than 100 µg/ml of DNA to limit non-specific amplification.

      Helpful?

  4. What are the possible reasons for problems when trying to review results for MP0035-1KT?

    1 answer

    1. Possible reasons for problems when trying to review results for MP0035-1KT could include incomplete removal of phenol during DNA preparation by phenol:chloroform extraction, leading to poor PCR results. It is also important to ensure the use of the recommended product, D9307, JumpStart™ Taq DNA Polymerase, as internal testing has indicated occasional sensitivity issues when using polymerases from other manufacturers. Additionally, the presence of inhibitors in the samples may affect the internal control band in the results. Lastly, mishandling of the vials and loss of lyophilized reagents could compromise the sensitivity of the assay.

      Helpful?

  5. Do mycoplasma-active antibiotics include pen/strep? Can media continue to be used with pen/strep prior to mycoplasma testing for kit R: MP0035?

    1 answer

    1. Pen/strep can be used with the cell culture before assaying with this kit. Pen/strep does not affect mycoplasma, nor does it affect the sensitivity of this kit.

      Helpful?

  6. What are the components of the rehydration buffer for MP0035? Can the rehydration buffer M4694 be substituted by another buffer?

    1 answer

    1. The rehydration buffer M4694 in Product No. MP0025 cannot be substituted by the PCR buffer P9367, and vice versa. The composition of the PCR 10X reaction buffer P9367 in the MP0025 kit is 100 mM Tris-Hcl, pH 8.5, 750 mM KCl, 30 mM MgCl2. Similarly, the replacement of Rehydration Buffer (Catalog Number M4694) with 10X PCR Buffer, Catalog No. P2192 is not possible. The composition of the 10× buffer P2192 is 100 mM Tris-HCl, pH 8.3 at 25°C; 500 mM KCl; 15 mM MgCl2; 0.01% gelatin.

      Helpful?

  7. When should samples be heat inactivated for using MP0035, Mycoplasma PCR Detection Kit?

    1 answer

    1. The possibility that as the slides dry, salts from the PBS are evaporating and leaving a white residue or film is considered.

      Helpful?

  8. What is the difference between product no. MP0040A (LookOut® Mycoplasma qPCR Detection Kit) and MP0035???

    1 answer

    1. The key differences between these kit are as follows:

      MP0035
      Standard PCR
      primers/dntps precoated on tubes
      Tubes included
      Taq Polymerase not included
      Requires 40 cycles at 30 seconds

      MP0040A
      qPCR only
      primers/dntps require addition to reaction
      Tubes not included
      Taq Polymerase included in primer/dntp mix
      Requires 45 cycles at 30 seconds

      MP0035 is suitable for 24 reactions while MP0040A is suitable for 25 reactions.

      Please see the datasheets for both products to see experimental differences in the procedures:
      MP0035
      https://www.sigmaaldrich.com/deepweb/assets/sigmaaldrich/product/documents/213/434/mp0035bul.pdf
      MP0040A
      https://www.sigmaaldrich.com/deepweb/assets/sigmaaldrich/product/documents/109/594/mp0040abul.pdf

      Helpful?

  9. I purchased the JumpStart Taq DNA Polymerase in addition to the kit. This also included a 10x PCR buffer. Is this also needed or only the rehydration buffer of the kit?

    1 answer

    1. This kit has been tested and optimized for use with JumpStart Taq DNA Polymerase, Catalog Number
      D9307. The Jumpstart Taq 10X buffer supplied with the Jumpstart Taq is not used in this kit reaction set up. The Jumpstart taq is used alone with the rehydration buffer provided with the kit.

      Helpful?

  10. Which species of Mycoplasma will Product MP0035, LookOut® Mycoplasma PCR Detection Kit, recognize?

    1 answer

    1. Detection of all mycoplasma species tested so far and usually encountered as contaminants in cell cultures  This kit will detect A. laidlawii, M. agalactiae, M. arginini , M. arthritidis, M. bovis, M. cloacale, M. falconis, M. faucium, M. fermentans, M. hominis, M. hyorhinis, M. hyosynoviae, M. opalescens, M. orale, M. primatum, M. pulmonis, M. salivarium, M. spermatophilum, M. timone.  This kit is not suitable for detection of M. pneumoniae, U. urealyticum, or other clinically associated species.

      Helpful?

1–10 of 16 Questions  

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