polymerase chain reaction (PCR)

Specimen

• 10 ng of DNA is adequate for quantitative PCR measurements
• crude cell lysate, serum & whole blood may be used directly
• avoid anticoagulants
  • heparin is a strong inhibitor of PCR
  • citrate & EDTA are acceptable anticoagulants
• heme is a strong inhibitor of PCR
• specimens should be stored at -20 degrees C or below

Procedure

• Amplification of DNA through action of cycles of denaturation of DNA, annealing of primers to DNA, & DNA synthesis using thermostable DNA polymerases.
• Theoretically, for n cycles of replication, the DNA may be amplified 2(exp)n fold.
• Careful selection of primers complementary to unique DNA sequences on the amplified DNA is necessary.

Interferences

• common, including
  • carry-over exogenous DNA from contamination of glassware
  • DNA on skin
  • aerosolized DNA

Negative controls are of critical importance.

Notes

• Turnaround time: 4-6 hours

More general terms

• special chemistry test

More specific terms

• multiplex polymerase chain reaction (PCR) assay
• reverse transcriptase polymerase chain reaction (RT-PCR)
• telomeric repeat amplification protocol (TRAP)

Additional terms

• molecular diagnostic test