First trimester prenatal diagnosis of trisomy 21 in fetal cells from maternal blood.
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Adult↗
Biomedical subjects
S Wachtel
Publications and source records attributed to S Wachtel.
Mutations in the conserved domain of SRY are uncommon in XY gonadal dysgenesis.
In order to evaluate the role of SRY in the determination of the testis, we sequenced the conserved domain of the SRY gene in 8 patients with 46,XY gonadal dysgenesis and 3 patients with related disorders, and compared our data with those obtained in 6 other similar studies. In our study, a 609-bp fragment of SRY was amplified by the polymerase chain reaction and the internal conserved motif was sequenced. SRY sequences did not differ from those in normal males in any of our patients. Overall, 5 de novo mutations have been identified among 56 patients with sporadic XY gonadal dysgenesis (8.9%), and 2 de novo mutations have been identified among 18 patients with related conditions (11%). The unexpectedly low frequency of mutations within the SRY conserved domain in these patients could be caused by undetected Y-linked mutations outside the conserved domain in regions that control transcription during development (e.g., promoter/enhancer regions) or to downstream mutations in other sex-determining genes that need not map to the Y.
Adolescent↗
Fetal cells in the maternal circulation: isolation by multiparameter flow cytometry and confirmation by polymerase chain reaction.
During pregnancy, nucleated fetal erythrocytes enter the maternal circulation and can be isolated efficiently from the maternal cells by multiparameter flow cytometry. Male DNA, implying presence of a male fetus, can be identified in flow-sorted maternal blood by polymerase chain reaction with oligonucleotide primers flanking single-copy Y-specific DNA sequences. Among flow-sorted samples, we correctly identified fetal sex in 17/18 (94%) pregnancies of 10-21 weeks gestation. Maternal blood thus provides a potential opportunity for prenatal diagnosis that could preclude the need for invasive procedures in current use.
Base Sequence↗
DNA fingerprinting in reptiles: Bkm hybridization patterns in Crocodilia and Chelonia.
The simple tetranucleotide repeat GATA (GACA) occurs in all eukaryotes so far studied. In many species, the arrangement of these sequences varies considerably among individuals. Thus GATA (GACA) type probes produce DNA fingerprints when hybridized with restricted DNA from different individuals within a species. Banded krait minor (Bkm) satellite DNA (related to sequences originally recovered from the W chromosome of the banded krait and consisting essentially of a series of GATA repeats) is found in a wide spectrum of vertebrates and invertebrates. We used the Bkm 2(8) clone to evaluate the occurrence of this satellite in DNA from five species of Crocodilia and six species of Chelonia, including the sea turtles Chelonia mydas and Lepidochelys kempi. Well-resolved DNA fingerprints were obtained. Among the crocodilians, fewer restriction fragments were generated and fewer of the fragments were polymorphic, than among the chelonians, consistent with the view that the crocodilians are less divergent within species. The Bkm 2(8) clone can accordingly be used to advantage in individual, familial, and population studies, and perhaps in the evaluation of taxonomic relationships in these animals. This is of potential value in endangered species such as C. mydas and L. kempi.
Alligators and Crocodiles↗
Bkm satellite DNA and ZFY in the coral reef fish Anthias squamipinnis.
We studied DNA from the protogynous sex-changing fish Anthias squamipinnis to evaluate the recent observation that male-specific bands are identified after hybridization with Bkm, a probe originating in the W chromosome of the snake Bungarus fasciatus. Sex-specific hybridization would imply modification of DNA structure during the sex-changing process. No sex-specific Bkm fragments were identified in our study, after digestion of DNA from 15 males and 11 adult females, despite the use of 12 different restriction enzymes. However, hybridization with Bkm did produce a distinct fingerprint pattern, similar to the fingerprint patterns described for other species after hybridization with GATA (GACA) type probes. In other experiments, the pDP1007 probe, which identifies the ZFY gene in the male-determining region of the human Y chromosome, generated identical hybridization patterns in DNA from males and females of A. squamipinnis and estimation of DNA mass by flow cytometry revealed identical genome sizes.
Animals↗
Sex-specific DNA in reptiles with temperature sex determination.
Banded krait minor ("Bkm") satellite DNA, originating in the W-chromosome of the snake Bungarus fasciatus, has been found in the genome of diverse eukaryotic species including fruit fly, quail, and horse. Concentrations of Bkm have been found in the presumptive W-chromosome of snakes with isomorphic sex chromosomes and in the male-determining region of the Y-chromosome in mouse and man. We therefore asked whether Bkm-related DNA might be present in quantitative excess in DNA from males or females in two related species of sea turtle, Chelonia mydas, in which sex is determined by the temperature of the incubating egg, and Lepidochelys kempi, in which the critical sex-determining temperature has recently been described. Filter hybridization with the Bkm 2(8) probe revealed male-specific fragments in both species; female-specific fragments were also revealed in C. mydas. Sex-specific DNA sequences in temperature-sex-determined species such as Kemp's ridley and the green turtle were unexpected, but could be explained if there were an underlying genetic mode of sex determination in these animals, or alternatively, if temperature-influenced sex determination involved structural modifications in DNA adjacent to, or directly concerned with, the sex-determining genes. If these results are confirmed across a broader sample of sea turtles, the techniques described in this paper might be used routinely to identify gener in the young of these endangered animals, in which male and female are grossly indistinguishable.
Animals↗
Direct estimation of serological H-Y antigen by flow cytometry.
This study was undertaken to evaluate the efficiency of flow cytometry in the detection of the serological H-Y antigen, and to survey expression of H-Y in the normal human population. Peripheral blood leukocytes (granulocytes) were reacted with monoclonal H-Y antibody, gw-16, and with FITC-conjugated goat anti-mouse IgG, and then were scored for fluorescence in the flow cytometer. Assigned H-Y phenotype was correlated accurately with sex phenotype in 33 out of 38 women and 62 out of 65 men. Data were evaluated by exploratory data analysis (EDA).
Antibodies, Monoclonal↗
Diacetyl-splenopentin (BCH 069) did not affect the concentrations of various hormones in men.
We investigated the effect of BCH 069 (similar structure as thymopentin) on concentrations of hormones of the hypophysis, adrenals, gonads and thyroid glands in men after a therapy period of 4 or 6 weeks. No significant effects of BCH 069 three times weekly 50 mg s.c. for 4 weeks or i.v. for 6 weeks on serum concentrations of prolactin, LH, TSH, cortisol, testosterone, progesterone, estradiol and ETR were observed.
Adult↗
Relative deoxyribonucleic acid content of interphase leukocytes by flow cytometry: a method for indirect diagnosis of chromosomal abnormalities with potential for prenatal diagnosis.
A major goal of prenatal cytogenetic analysis is the development of minimally invasive techniques by which all pregnancies may be screened. That nucleated fetal cells exist in the maternal circulation raises the possibility that their cytogenetic status could be determined. However, these cells may not respond to commonly used mitogens. Thus it will probably be necessary to develop methods to analyze nondividing (interphase) cells. We therefore evaluated the use of flow cytometry as a means of determining whether the relative deoxyribonucleic acid content of G0-G1 leukocytes (expressed as the deoxyribonucleic acid index) could be used to verify aneuploidy or other chromosomal abnormalities. Our findings indicate this is indeed possible. We determined that the mean deoxyribonucleic acid index of circulating leukocytes from normal adult men (n = 15) was significantly different from that of leukocytes from normal adult women (n = 15). Similar results were obtained in leukocytes from umbilical cord blood of normal male neonates (n = 15) and normal female neonates (n = 15). Most importantly, values for leukocytes from each of 13 aneuploid individuals fell outside the range of values for leukocytes from normal adults of the same sex.
Adult↗
Purification and analysis of a sex-specific antibody.
Analysis of the male-specific H-Y antigen is difficult, in part, because of the tendency of "male-specific" antisera to bind cell surface components found in male and in female cells. Some insight as to the nature of that difficulty is provided by biochemical evaluation of H-Y antibody. In this study, purification of a monoclonal H-Y antibody with ammonium sulfate or Protein-A Sepharose, revealed the possibility of microheterogeneity. Despite evidence of multiple subtypes, Protein-A elution profiles suggested that the male-specific activity of the antibody resided in an IgG2a moiety. This was borne out by decreased activity after absorption of the IgG2a subtype with male cells, and by reaction of the monoclonal antibody with mouse subtype-specific antisera in an ELISA. Combined analysis using biological (absorption), biochemical (EF and PAGE) and immunological (ELISA) methods could find applicability in other complex systems.
Animals↗
DNA analysis by flow cytometry.
Accurate quantification of DNA from cells of several species is possible with flow cytometry. When one species is used as a reference, cytometric readings from two or more different species can be compared to obtain relative percent DNA or DNA indices. Differences in DNA from the male and female of the same species also can be measured. The method allows rapid screening of chromosomal abnormalities among large clinical populations, and evaluation of errors of sex determination such as XY sex reversal.
Animals↗
Sex selection with monoclonal H-Y antibody.
H-Y antigen defined by antibody from male-sensitized female mice has been reported in male embryos of mouse, rat, cattle, goat, pig, and sheep. We now describe the use of monoclonal H-Y antibodies in identification of male and female during embryo transfer in cattle. Monoclonal H-Y antibodies were applied with fluorescein-isothiocyanate-conjugated goat antimouse immunoglobulin (Ig), and the embryos were scored for fluorescence under ultraviolet light. In alternative trials with 149 embryos, the method could be applied with 73% to 82% efficiency. Pregnancy rates for treated embryos were at least as high as those expected for untreated embryos. The calves that developed from the treated embryos are normal and growing.
Animals↗
Reaction of monoclonal H-Y antibody in the ELISA.
Monoclonal H-Y antibody of demonstrated specificity was reacted with soluble H-Y antigen in a newly-developed enzyme-linked immunosorbent assay (ELISA). Typing of cell lines was accomplished by qualitative and quantitative absorption. In one case, the antibody was subdivided into equal portions, each of which was absorbed with 1, 3, 6 or 12 X 10(6) cells from male or female; after absorption, the cells were discarded and the antibody was titrated for residual activity in the ELISA. With increasing cell numbers, optical density scores for male-absorbed antibody approached baseline levels. Although male-female differences were pronounced and highly significant, there was also a fall in reactivity after absorption with female cells.
Animals↗
The endocrinologic evaluation of a 45,X true hermaphrodite.
A 13-year-old 45,X girl presented with spontaneous menses and the acute onset of progressive hirsutism. Complete endocrinologic evaluation of hirsutism was unremarkable. An exploratory laparotomy revealed a gonad containing ovarian and testicular tissue. Chromosome analysis from four different tissues confirmed the karyotype to be 45,X. She was H-Y antigen positive.
Adolescent↗
Enzyme immunoassay of H-Y antigen: experimental and clinical applications.
A newly developed enzyme-linked immunosorbent assay (ELISA) was applied for studying H-Y antigen in buffaloes, cattle, horses and humans. A monoclonal H-Y antibody was absorbed with cells from males or females and was then tested against fluid samples known to contain soluble H-Y antigen. In this system, positive absorption manifested itself by a fall in optical density relative to the optical density scored using unabsorbed antibody; this finding signified the presence of H-Y in the absorbing cells. In each of the four species, the fall in optical density was pronounced after absorption with male cells, but some decrease was also evident after absorption with female cells, indicating a degree of nonspecific attachment of the antibody.
Animals↗
On the expression of H-Y antigen in transsexuals.
Histocompatibility-Y (H-Y) antigen, the presumptive inducer of the mammalian testis, is present in the cells of normal males and not in the cells of normal females. Recent reports have implied that patients with transsexualism exhibit H-Y antigen phenotypes at variance with those of normal males and females and, thus, that H-Y serology might provide a tool for the diagnosis and study of the transsexual condition. We therefore evaluated blood and testicular cells from 21 male-to-female transsexuals using conventional and monoclonal H-Y antibodies. We found no evidence of abnormal H-Y phenotype. Five of the patients were interviewed postoperatively by two examiners and rated for the diagnosis of transsexualism. Three of the five were rated primary transsexual by one or both examiners, and two were rated secondary transsexual.
Adult↗
On the study and control of sex differentiation.
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Animals↗
H-Y antigen in a 46,XX female with dysgenetic ovaries.
Hypogonadism secondary to ovarian dysgenesis or resistant ovary syndrome was diagnosed in a 19-yr-old obese woman with primary amenorrhea, a 46,XX karyotype, and an H-Y+ cellular phenotype. Small ovoid gonads (1.5 X 0.6 cm) were found found bilaterally; these were encased in a dense venous network. The stroma was ovarian, and primordial follicles and some primary follicles were present, but there were no follicles at or beyond the antrum stage. There was no evidence of testicular tissue and no evidence of malignancy. Analysis of serological data indicated the possibility of residual H-Y antigen in the blood cells of the mother.
Adult↗