Microsatellite analysis provides efficient confirmation of fetal trophoblast isolation from maternal circulation

Fetal trophoblasts can be found in maternal circulation from an early stage of pregnancy and thus provide a potential source of DNA for non-invasive prenatal diagnosis. We have developed a two-step method for trophoblast isolation between the 8th and 12th week of pregnancy. Blood was sampled from 14 women undergoing termination of pregnancy or spontaneous abortion. Immunomagnetic beads precoated with HLA class I and II, and with anti-cytokeratin-18 monoclonal antibodies, were used to remove CD8+ and other maternal cells, and to select for fetal trophoblasts, respectively. Microsatellite analysis was performed on DNA extracted from the isolated, maternal, paternal and placental cells after PCR amplification. Recovery of the trophoblasts was confirmed in 13/14 cases (93%) by the identification of an identical microsatellite pattern for fetal and placental cells. Further evidence was the presence of heterozygous alleles of both maternal and paternal origin. The correct prediction of gender in all five male fetuses was an additional confirmation of trophoblast recovery. We conclude that trophoblasts can be effectively isolated from maternal blood in the first trimester, and by using polymorphic microsatellite markers to confirm sample purity, this method has potential future application in prenatal diagnosis. Copyright © 2001 John Wiley & Sons, Ltd.     

Pseudomosaic centric fission of chromosome 4 in amniotic cells

This paper describes a case of pseudomosaic centric fission of chromosome 4 detected in amniotic fluid cell culture. The pregnancy went to term and the newborn had a normal chromosomal constitution.     

Decolorization of reactive Brilliant Blue KN-R by immobilized cells of Aspergillus ficuum

Aspergillus ficuum was immobilized with sodium alginate, and decolorization of Reactive Brilliant Blue KN-R was studied on immobilized and free Aspergillus ficuum. The optimal preparation condition of the strain immobilization was obtained by the orthogonal test, it is sodium alginate 3%, CaCl2 5%, wet mycelia 30 g/L, calcific time 8 h. It was found that the immobilized cells could effectively decolorize Reactive Brilliant Blue KN-R, the optimum temperature and pH were 33℃ and 5.0, respectively. The kinetics study of decolorization of immobilized cells showed that the decolorization of Aspergillus ficuum immobilized conformed to zero-order reaction model. The decolorization efficiency of immobilized cell compared with that of free cell in different physical conditions. Results showed that the decolorization of immobilized cells with mycelia had the best efficiency. The immobilized cells could be reused after the first decolorization.     

Maternal cell contamination of amniotic fluid cell cultures from two consecutive pregnancies complicated by fibroids

The detection of maternal cells in amniocyte cultures is thought to be due to the outgrowth of cells from small fragments of maternal tissue removed by the amniocentesis needle. An unusual case is reported in which maternal cell contamination (MCC) was found in the cell cultures from a woman in two different amniocenteses from two consecutive pregnancies. Both pregnancies were complicated by the presence of fibroids and the fibroid tissue may have been the source of the maternal cells. A history of an amniocentesis in which there was MCC of cell cultures, or the detection of fibroids, may pose an additional risk for MCC attributable misdiagnosis in prenatal genetic studies.     

Improved direct molecular diagnosis and rapid fetal sexing

Adaptations of the techniques of modern molecular biology to prenatal diagnosis has opened new avenues for the detection of genetic diseases. We have taken advantage of the rapid adhesion of colony forming cells in cultured amniotic fluid samples to develop an improved method for molecular diagnosis. By employing the cell adherence regime sickle cell diagnosis using Mst II can be undertaken directly. In addition, hybridization with a cloned repetitive sequence that is of Y origin and has limited autosomal homology permits rapid fetal sexing in 3 to 4 days without compromising conventional cytogenetic or biochemical analysis. This combination of techniques provides a useful adjunct to convential prenatal genetic diagnosis in the second trimester.     

Maternal cell contamination in cultured chorionic villi: Comparison of chromosome Q-polymorphisms derived from villi,fetal skin,and maternal lymphocytes

Maternal cell contamination of chorionic villi (CV) samples used for first trimester prenatal diagnosis can cause obvious and/or unrecognized diagnostic dilemmas. The purpose of this investigation is to assess the frequency of maternal cell contamination (MCC) in chorionic villus samples and to evaluate selected parameters which might predict where contamination is more likely to have occurred. Maternal lymphocytes, chorionic villi from ultrasonically directed transcervical catheter aspiration, and fetal tissue were obtained at 8–11 weeks gestation from 45 patients undergoing elective termination. Quinacrine (Q) banded metaphases were compared from duplicate direct preparations of chorionic villi; cultured chorionic villi, fetal fibroblast tissue cultures, and maternal lymphocyte cultures. Q-polymorphisms in metaphase chromosomes were 100 per cent concordant between fetal tissue and direct CV preparation. However, evidence for maternal cell contamination occurred in 13.1 per cent of cultured chorionic villi preparations where polymorphisms were found to be identical between maternal and cultured CV and both distinct from fetal tissue preparations. Where MCC was identified, it was noted that CV cell cultivation interval was prolonged (24.2±6.8 days) compared with non-contaminated cultures (14.1±4.4 days) (p <0.05). We conclude that maternal cell contamination is a significant problem with chorionic villus sampling. Where direct preparations are not employed or when cultures are ‘slow growing’, MCC may be a significant and unrecognized complication re: fetal diagnosis. Direct preparations, multiple cultures, quinacrine banding, and maternal Q-polymorphism comparisons can minimize diagnostic dilemmas secondary to maternal cell contamination. Q-polymorphism comparisons between maternal and fetal chromosomes should be included in all instances where cultured chorionic villi are utilized for fetal diagnosis and where direct preparations are not available.     

Correct prenatal diagnosis of a hurler fetus where amniotic fluid cell cultures were of maternal origin

Contamination of amniotic fluid cell cultures by maternal cells can be expected to lead to misdiagnosis of fetal genotype in 0·1 to 0·5/100 cultures, when assays are carried out directly on cultured cells. Chemical analysis of the cell-free amniotic fluid supernatant may overcome this source of error and has the added advantages of speed and independence from amniotic cell culture failure. We describe a pregnancy at risk for Hurler's disease where amniotic cells cultured at amniocentesis had a female karyotype and an α-iduronidase activity towards both phenyl and 4-methylumbelliferyl substrates at the lower end of the normal range, suggesting a heterozygous fetus. An affected fetus was predicted, however, because of a high concentration of dermatan sulphate in the amniotic fluid. The discrepancy between these findings was shown to be due to maternal cell contamination of amniotic fluid cell cultures by the birth of a male infant with Hurler's disease.     

固定化细胞技术处理含酚废水的研究

固定化细胞技术的关键是所用载体材料的性能。本文力求寻找一种价格低、寿命长、效率高的载体。通过同一菌种在固定状态和游离状态降解含酚废水的实验对比，证明红砖碎粒是一种优良的载体材料。利用正交实验，确定了该菌种在固定时的最佳运行条件。并对游离细胞和固定细胞降解苯酚的过程进行了动力学分析。结果表明，在两种情况下，该菌种降解苯酚的过程均符合Ｍｏｎｏｄ模型。     

一种水质污染生物监测的新方法——蝌蚪肠细胞染色体畸变分析

发展了一种水生脊椎动物蝌蚪肠细胞染色体畸变的分析方法,可作为诱变剂污染水源的早期报警系统。该方法的优点:1)灵敏度高;2)蝌蚪在我国分布广、数量多、捕捉方便、费用低、易饲养管理。尤其是蝌蚪的核型由22条大的中部和亚中部着丝点染色体所组成,适于毒理遗传学研究;3)蝌蚪具有转化前诱变剂或前致癌剂为活性成份的微粒体活化系统。