TY - JOUR
T1 - Resistance to activated protein C
T2 - Comparison of three different PCR methods for detection of FV R506Q
AU - Voelkerding, Karl V.
AU - Huber, Suzanne
AU - Strobl, Frank
AU - Wu, Lambert A.
AU - Sabatini, Linda M.
AU - Anderson, Mary
AU - Lutz, Charles T.
PY - 1996
Y1 - 1996
N2 - Background: Resistance to activated protein C (APC) is the most prevalent identifiable risk factor for inherited thrombophilia. Over 90% of APC resistance results from a single point mutation in the Factor V gene. The mutation, termed FV R506Q or FV Leiden, predicts an abnormal Factor Va protein in which arginine, at amino acid position 506, is replaced by glutamine, rendering Factor Va resistant to proteolytic inactivation by APC, thus establishing a life long hypercoagulable state. The current study compared three different polymerase chain reaction (PCR)-based approaches for the detection of FV R506Q. Methods and Results: Sixty-seven patient blood samples were genotyped by (1) analyzing for loss of a Mill I recognition site, an acquired restriction-fragment-length polymorphism (RFLP) due to FV R506Q; (2) primer-engineered RFLP wherein the presence of FV R506Q results in generation of a novel Nla III recognition site; and (3) allele-specific PCR. Sixty-five of 67 patient samples yielded concordant genotype results by all three PCR methods. Of the remaining 2 of the 67 patients, a "nondiagnostic" result was obtained for either allele-specific PCR or primer-engineered RFLP. Conclusions: A comparative analysis of 67 patient samples demonstrated that primer engineered RFLP and allele-specific PCR offer feasible alternative or confirmatory testing approaches to Mnl I RFLP for the detection of FV R506Q. A high degree of diagnostic concordance was observed for all three methods, and no false positive or negative results were observed with the Mnl I RFLP technique.
AB - Background: Resistance to activated protein C (APC) is the most prevalent identifiable risk factor for inherited thrombophilia. Over 90% of APC resistance results from a single point mutation in the Factor V gene. The mutation, termed FV R506Q or FV Leiden, predicts an abnormal Factor Va protein in which arginine, at amino acid position 506, is replaced by glutamine, rendering Factor Va resistant to proteolytic inactivation by APC, thus establishing a life long hypercoagulable state. The current study compared three different polymerase chain reaction (PCR)-based approaches for the detection of FV R506Q. Methods and Results: Sixty-seven patient blood samples were genotyped by (1) analyzing for loss of a Mill I recognition site, an acquired restriction-fragment-length polymorphism (RFLP) due to FV R506Q; (2) primer-engineered RFLP wherein the presence of FV R506Q results in generation of a novel Nla III recognition site; and (3) allele-specific PCR. Sixty-five of 67 patient samples yielded concordant genotype results by all three PCR methods. Of the remaining 2 of the 67 patients, a "nondiagnostic" result was obtained for either allele-specific PCR or primer-engineered RFLP. Conclusions: A comparative analysis of 67 patient samples demonstrated that primer engineered RFLP and allele-specific PCR offer feasible alternative or confirmatory testing approaches to Mnl I RFLP for the detection of FV R506Q. A high degree of diagnostic concordance was observed for all three methods, and no false positive or negative results were observed with the Mnl I RFLP technique.
KW - Coagulation
KW - Factor V leiden
KW - Thrombosis
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U2 - 10.1016/S1084-8592(96)70012-7
DO - 10.1016/S1084-8592(96)70012-7
M3 - Article
AN - SCOPUS:0000027837
SN - 1084-8592
VL - 1
SP - 297
EP - 304
JO - Molecular Diagnosis
JF - Molecular Diagnosis
IS - 4
ER -