| Co-Investigator(Kenkyū-buntansha) |
OYAMA Rio Iwate Medical University, School of Medicine, Research Associate, 医学部, 助手 (20291619)
SHOJI Thdahiro Iwate Medical University, School of Medicine, Research Associate, 医学部, 助手 (00337148)
SUGLYAMA Thni Iwate Medical University, School of Medicine, Professor, 医学部, 教授 (40162903)
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| Budget Amount *help |
¥3,700,000 (Direct Cost: ¥3,700,000)
Fiscal Year 2004: ¥500,000 (Direct Cost: ¥500,000)
Fiscal Year 2003: ¥500,000 (Direct Cost: ¥500,000)
Fiscal Year 2002: ¥2,700,000 (Direct Cost: ¥2,700,000)
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| Research Abstract |
The object of the present study was to attempt to apply Laser Scanning Cytometer (LSC) to automated detection of fetal nucleated red blood cells (FNRBC) in maternal peripheral blood. 1) Fetal cell isolation : We investigate the effectiveness of a relatively easy and efficient method of sorting fetal nucleated red blood cells (FNRBCs) from maternal peripheral blood, particularly during early gestation periods, by a combination of specific gravity centrifugation and magnetic cell sorter (MACS). Without prior knowledge of the sex of each fetus, we determined gender by analyzing a Y-chromosome specific sequence by nested-PCR, using 10 ml of the peripheral blood of healthy primigravida women at different stages of gestation (first trimester: n17, second trimester : n=13, and third trimester : n=19). The results of this prenatal sex determination were compared to the sex of newborns. The sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) of the pres
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ent method during the first trimester was 81.8%, 100%, 100% and 75%, respectively; during the second trimester, 50%, 80%, 80%, and 50%, respectively; and during the third trimester, 63.6%, 25%, 53.8% and 33.3%, respectively. 2) LSC technique: A venous blood sample (lOmI) was collected from 29 women carrying a male fetus between 5 and 38 weeks of gestation who gave informed consent to participate in the present study. Twenty-four of the blood samples were taken from normal pregnant woman, while 5 were taken from women hospitalized to inhibit labor. Eukaryotic cells were promptly separated by gravity centrifugation. Then, cells negative for CD45 monoclonal antibody and positive for glycophorin A monoclonal antibody (GA) were selectively collected using magnetic-activated cell sorter (MACS) to obtain FNRBC. FNRBCs were determined by fluorescence in situ hybridization (FISH) Y specific probes. FNRBCs were found with an average count of 9.7±4.2 per 10 ml of venous blood. We found no relationship between stage of pregnancy and number of FNRBC. There was a good correlation between results of manual and automated screening for the 29 samples examined (R2= 0.843). These results show this prenatal sex determination method has a highly accurate diagnostic rate during the first trimester, suggesting that it could be developed as a practical, non-invasive prenatal diagnostic technique for use during early gestation periods. Less
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