Abstract
A study was conducted on simple methods for the prenatal diagnosis of hypoxanthine-guanine phosphoribosyltransferase (HPRT) deficiency using amniotic cells. The amniotic cells were cultured in triplicate in 1 ml of MEM medium containing 10% fetal calf serum, antibiotics and "C-hypoxanthine at 37°C in a humidified atmosphere of 5% CO, and 95% air using sterile microplates with 24 flat-bottom wells. Following appropriate time of culture, the plates were washed with cold physiological saline, and the radioactivities of "C-hypoxanthine in the cells suspended in the trypsin solution were counted. Firstly, the influence of cell number and culture time on the uptake of "C-hypoxanthine in normal cells was studied. In the case of normal cells, it was shown that only 5,000 cells cultured for 12hours was sufficient to clarify the incorporation of "C-hypoxanthine. Next, batches of 10,000 amniotic cells from a normal subject and from two cases at risk for HPRT deficiency were cultured with "C-hypoxanthine and or "C-adenine for 24 hours. Marked uptakes of "C- hypoxanthine and "C- adenine were shown by cells from both thenormal subject and those from one patient. However, in the cells from the patient whose cell lysate did not show any HPRT activity, uptake of "Cadenine in the cells was evident while that of "C-hypoxanthine was not. Thus, this method seems to demonstrate simple and accurate diagnosis of HPRT deficiency in thefetus.
