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Quality Assurance/Quality Control Manual: Ohio Water Microbiology Laboratory

Thermal Cyclers

The OWML uses two real-time thermal cyclers used to perform the quantitative polymerase chain reaction (qPCR), both are from Applied Biosystems: The AB 7500 Real Time PCR System (AB 7500) and the AB StepOnePlus Real Time PCR System. qPCR is done to amplify microbial DNA targets through a series of temperatures changes. Numerous projects use qPCR for a variety of applications including microbial source tracking, detection and/or quantification of enteric viruses, and detection and/or quantification of bacterial indicators.

Quality assurance measures for both thermal cyclers are as follows:

  • Background calibration is performed monthly.
  • A pure-dye spectral assay is done annually or as needed (if replicate amplification curves are continually wavy or non-uniform).
  • A RNase P verification run is done as needed (as a troubleshooting measure).
  • A regions of interest (ROI) plate is run as needed (following lamp replacement or if the system is jostled).
  • Annual preventative maintenance is done annually by AB technicians.
  • All quality assurance checks are recorded in the LIMS.