OBJECTIVETo establish an effective laboratory examination system for carrier detection and prenatal diagnosis of haemophilia A (HA).

METHODSTwenty-five carriers of severe HA were directly detected by long-distance PCR (LD-PCR) in search of the factor FVIII (FVIII) gene inversion. Prenatal diagnosis was carried out using pregnant woman's venous blood sample, husband's venous blood sample and fetal navel venous sample at 20-24 weeks of gestation. The plasma coagulation factor VIII activity (FVIII:C) was detected by one-stage method. The concentration of von Willbrand factor (Vwf) was assayed by ELISA. Prenatal diagnosis was finally made by LD-PCR. The results of LD-PCR were proved by DNA sequencing.

RESULTSEight out of 25 cases were diagnosed as having FVIII geneinversion. Four of these 8 carriers underwent the LD-PCR for prenatal diagnosis, and 2 of them had to terminate pregnancy because their fetuses were diagnosed as having HA. The other two carriers were finally diagnosed to have normal fetuses by combined use of LD-PCR with plasma FVIII:C, vWF in pregnant woman's venous blood, husband's venous blood and fetal navel venous blood, and the one-year follow-up study demonstrated that the babies were normal and living well.

CONCLUSIONLD-PCR technique was adopted in this study to detect the factor VIII gene inversion; it could accurately and rapidly diagnose the severe cases of HA and could be used for the HA carriers in need of pregnant diagnosis.

Factor VIII ; genetics ; Female ; Hemophilia A ; diagnosis ; genetics ; Humans ; Polymerase Chain Reaction ; methods ; Pregnancy ; Prenatal Diagnosis ; methods ; Reproducibility of Results

The aim of current study was to detect intron 22 inversion of factor VIII gene in severe hemophilia A (HA) patients and screen the carriers of the gene inversion. Fifty-five cases of severe HA were involved and factor VIII gene inversion was detected and identified by long distance-PCR (LD-PCR) and 0.6% agarose gel electrophoresis. The 11 kb and 12 kb bands indicate the factor VIII gene inversion and non-inversion, respectively. Occurring of both 11 kb and 12 kb bands indicates a carrier of the inversion. The results showed that factor VIII gene inversion existed in 22 out of 55 cases, which accounted for about 40% of total detected patients. Five carriers of factor VIII gene inversion were diagnosed from the members in 15 families. In conclusion, LD-PCR assay is a simple, rapid and accurate method for detection of factor VIII gene inversion, and this approach is helpful in screening, carrier testing, and prenatal diagnosis of severe hemophilia A.
Adolescent ; Adult ; Antigens ; analysis ; Child ; Child, Preschool ; Chromosome Inversion ; Factor VIII ; genetics ; Hemophilia A ; blood ; genetics ; Humans ; Infant ; Male ; Polymerase Chain Reaction ; methods ; von Willebrand Factor ; immunology