Application Note: How to Pipette PCR Master Mix for Increased Accuracy in qPCR Results

Sensitivity of Polymerase Chain Reaction (PCR) based assays is one of the most important reasons for their success  and abundant use in scientific laboratories – particularly those working with clinical diagnostics and biopharmaceuticals. However, factors like pipetting technique can influence PCR assay results.

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Commonly Asked Questions  

  • How can pipetting affect qPCR results?  - Because PCR Master Mix contains proteins, improper pipetting can lead to the creation of bubbles in the sample, which can skew the results of the experiment. 
  • Which pipetting technique is best when using PCR Master Mix? - According to ISO guidelines, it is recommended to pre-wet the pipette tip 3 to 5 times, it is also recommended to use reverse pipetting technique as PCR Master Mixes are viscous. 
  • Is pipette calibration important when using PCR Master Mix? - There is a propensity for error when dealing with viscous materials such as PCR Master Mix. Therefore, it is important to eliminate as many risk factors as possible. Ensuring your pipette is regularly maintained and calibrated is an effective way to keep results accurate and reproducible.
  • Should I use a multichannel pipette for preparation of qPCR assays? - Using a multichannel pipette for qPCR assays ensures equal amounts of PCR Master Mix will be evenly and quickly distributed into several wells at once. This is a time-saving factor especially when working with a 96 or 384 well plate. Reproducibility can be increased in experiments further by using an electronic pipette and multi-dispensing mode.

CR Master Mix is often used during qPCR setup but can be challenging to pipette accurately. For in-depth information on best pipetting practices for PCR Master Mix, download this application note and learn about:  

  • Guidelines for pipetting viscous liquids
  • Benefits of electronic pipette use
  • Forward vs. reverse pipetting technique for optimal results
  • Best pipette tips to use for qPCR setup

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