DNA-based assay formats for detecting thrombosis factor V genotypes

Substitution at factor V nucleotide 1691, the Leiden mutation, is the most common known genetic cause of venous thrombosis. DNA-based assays for the detection of factor V Leiden alleles follow common strategies. The Amplify, Interrogate, and Detect strategy includes polymerase chain reaction (PCR) amplification of a factor V gene fragment, interrogation with a sequence-specific reagent, such as restriction endonuclease or oligonucleotide probe, and detection of the results of interrogation. The Differentially Amplify and Detect strategy uses sequence-specific PCR primers to selectively amplify factor V Leiden or normal alleles, followed by detection of the amplified fragments. Several assays have been validated in comparison studies and are reliable if performed properly. The choice of assay format depends upon laboratory experience and sample volume.