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Biomedical subjects

C M Thomas

Publications and source records attributed to C M Thomas.

At least 199 records · Page 11Linked to original sources
PubMed1990

A family of ATPases involved in active partitioning of diverse bacterial plasmids.

Low copy-number bacterial plasmids F (the classical Escherichia coli sex factor) and prophage P1 encode partitioning functions which may provide fundamental insights into the active processes which ensure that bacterial genomes are segregated to both daughter cells prior to cell division. These partitioning systems involve two proteins: ParA and ParB. We report that incC from the broad host-range plasmid RK2 is a member of the family of ParA partitioning proteins and that these proteins (as well as related proteins encoded by plasmids from Agrobacterium tumefaciens and Chlamydia trachomatis) contain type I nucleotide-binding motifs. Also, we show that the cell division inhibitor MinD is homologous to members of the ParA family. Sequence comparisons of ParB proteins suggest that they may contain sites for phosphorylation. We propose that ATP hydrolysis by the ParA protein may result in phosphorylation of the ParB protein, thereby causing a conformational shift necessary to separate paired plasmid molecules at the cell division plane.

Adenosine Triphosphatases↗
PubMed1990

The squamous cell carcinoma tumor marker in mammary carcinoma: comparison of polyclonal versus monoclonal antibody-based assays.

Pretreatment serum levels of squamous cell carcinoma (SCC)-antigen as determined with a polyclonal and monoclonal antibody-based assay were determined in 136 patients with invasive mammary carcinoma, 8 patients with ductal carcinoma in situ and 23 patients with benign breast diseases. For the polyclonal assay, the overall sensitivity was 44.8% at a cut-off level of 2.0 ng/ml and 25% for 2.5 ng/ml. For the monoclonal assay, the sensitivity was 11.0 and 25.0% at cut-off levels of 2.0 and 1.5 ng/ml, respectively. On the basis of analysis of specificity of the two assays for the benign control group, the cut-off level of the polyclonal assay has to be 2.5 ng/ml in the case of mammary carcinoma, of the monoclonal assay it has to be 2.0 ng/ml. For 175 serum samples of the whole group of patients (pre- and postoperative samples), the polyclonal RIA (x) and the monoclonal IRMA (y) were measured. The orthogonal regression analysis gave the equation: monoclonal IRMA = 1.03 polyclonal RIA - 1.15 ng/ml (Sy.x = 0.48; r = 0.7225). We also came to the conclusion that the SCC-antigen measured by either assay is different.

Antibodies, Monoclonal↗
PubMed1990

Postpartum return of pituitary and ovarian activity during lactation inhibition with the new dopamine agonist CV 205-502 and during normal lactation.

The new nonergot dopamine agonist CV 205-502 appears to be a promising alternative in the treatment of hyperprolactinemia. Regarding the potential use of CV 205-502 in infertility practice, we studied the influence of CV 205-502 on the return of endocrine fertility parameters during the physiological hyperprolactinemia of the puerperium. The resumption of pituitary and ovarian activity in 18 CV 205-502 treated women was compared with that in 10 bromocriptine-treated women. LH was measured by a new specific assay, which does not cross-react with hCG. This assay was also used in a second part of the study in which the pituitary function of 10 breast-feeding women was investigated. Both dopamine agonists adequately suppressed PRL. Pituitary secretion returned in the second week and was initially characterized by a high FSH/LH ratio. There were no major differences between CV 205-502 and bromocriptine. Ovulations occurred from day 18 on. The PRL rebound at the end of treatment seemed to play a role in the ovulation process. An acute increase of PRL just before midcycle was able to prevent ovulation. Breastfeeding women showed a delayed return of pituitary secretion: after a hypogonadotropic period, FSH returned in the third week and was followed by a period with a high FSH/LH ratio and follicular inactivity.

Aminoquinolines↗
PubMed1990

Human choriogonadotropin (hCG): comparisons between determinations of intact hCG, free hCG beta-subunit, and "total" hCG + beta in serum during the first half of high-risk pregnancy.

We have studied the concentrations of intact human choriogonadotropin (hCG) and the free hCG beta-subunit in blood samples from singleton pregnancies at risk for habitual or threatened abortion. The samples were obtained weekly between the 6th and 12th weeks and in the 14th and 16th weeks of gestational age. The concentrations of intact hCG, of the free hCG beta-subunit, and of "total" hCG (i.e., intact hCG and the free hCG beta-subunit: hCG + beta) were measured in serum by specific immunoassays. The distributional statistics (the 5th, 50th, and 95th percentiles) of "total" hCG + beta and of intact hCG showed very similar patterns, whereas the response curves for the free hCG beta-subunit showed very much lower serum concentrations. From these data we also estimated distributional statistics of the percent molar ratios of free hCG beta-subunit to intact hCG. We conclude that (a) the relatively small proportion of free hCG beta-subunit in serum during the first half of singleton pregnancy is far too low to interfere with the applied "total" hCG assay, as compared with the serum values obtained for intact hCG, and (b) the percent molar ratios of free hCG beta-subunit to intact hCG, or to "total" hCG + beta, never exceeded 1.0% throughout the period of pregnancy studied.

Abortion, Threatened↗
PubMed1990

Comparison of large volume culture to other methods for isolation of microorganisms from dialysate.

Patients on continuous ambulatory peritoneal dialysis (CAPD) who reside long distances from a CAPD center often use community medical laboratories to document and manage episodes of peritonitis. We examined the feasibility of using large volume cultures as an alternative to more costly and labor intensive methods and to enhance earlier recovery of microorganisms from these patients. Three methods of processing dialysate from patients on CAPD were compared: (a) inoculation of 400 mL dialysate into a transfer bag (Baxter Healthcare, Inc., Round Lake, IL) containing 100 mL of 5-fold concentrate of trypticase-soy broth: (b) inoculation of 5 mL into each of two Bactec bottles (Johnston Laboratories, Towson, MD): and (c) centrifugation of 50 mL and culture of the sediment without white cell lysis on plated media and two Bactec bottles. Of the 58 specimens cultured, 34 (59%) were positive by one or more methods. Antimicrobial activity was detected in 20/58 (34%) dialysates, which represent 54% of all no-growth cultures. Of the 34 culture-positive specimens, microorganisms were recovered on plated media in 22 (65%); by the centrifugation system in 32 (94%); by the routine Bactec system in 28 (82%); and by large volume culture in 30 (88%). The large volume culture system is an acceptable alternative to the more costly Bactec System and the labor intensive centrifugation method but does not significantly improve recovery of microorganisms.

Bacterial Infections↗
PubMed1990

Time-resolved fluoroimmunoassay for unconjugated estrogen in urine: comparison with a fluorometric assay for total estrogen and application in an in vitro fertilization program.

A time-resolved fluoroimmunoassay (TR-FIA) for unconjugated estrogens in human urine is described. 6-Keto-17 beta-estradiol-6-(O-carboxymethyl)oxime:bovine serum albumin is immobilized onto microtiter strip wells and the coated wells are incubated with 17 beta-estradiol standard preparations or unknowns with a polyclonal antiserum to 17 beta-estradiol-16,17-monosuccinyl:albumin. The antiserum-bound estrogen is detected by incubation with a europium-labeled anti-rabbit IgG that serves as both second antibody and tracer. After the immunoreactions, the bound portion of the labeled antiserum is quantified by dissociating the Eu3+ in a fluorescence-enhancement solution and measuring its fluorescence with a time-resolved fluorometer. The detection limit of the TR-FIA is 24 pmol of 17 beta-estradiol per liter; the analytical range extends to 1.8 nmol/L. This assay is a convenient alternative to radioimmunoassay and to the automated Kober-Ittrich fluorometry of total estrogen. Its advantages include short counting times; use of nonradioactive, stable reagents, all of which are commercially available; and more nearly complete automation. We conclude that this TR-FIA, compared with the Kober-Ittrich fluorometric assay (J Endocrinol 1957; 16:49-56), provides the clinician with equivalent information during follicular development therapy as part of an in vitro fertilization program.

Autoanalysis↗
PubMed1990

Heterophilic antibodies in human sera causing falsely increased results in the CA 125 immunofluorometric assay.

An in-house OC 125 monoclonal antibody-based "sandwich" immunofluorometric assay (IFMA) described previously (Clin Chem 1987; 33:2191-4) gave higher results for CA 125 in 37 of 123 serum samples than did a commercially available immunoradiometric assay (IRMA). Discordant results between the two assays became concordant when measurements of the samples were repeated with normal mouse serum (100 mL/L) included in the IFMA reagent. The murine immunoglobulins are thought to block the ability of the heterophilic antibodies in the human serum samples to cross-link the labeled antibody with the solid-phase antibody. Using an enzyme immunoassay, we demonstrated human anti-mouse antibodies (HAMA) in most of the discordant samples examined. We tested the ability of nonimmune sera from other animal species to lower the apparent CA 125 concentrations of the spurious samples and observed that rat, goat, and sheep serum were less effective than mouse serum. One serum sample was discovered to give a falsely increased CA 125 result with the IRMA, but this increase could be prevented by adding murine serum to the IRMA reagent. We conclude that falsely increased CA 125 results are best prevented by adding murine serum (or murine antibodies) to the assay buffer.

Antibodies, Heterophile↗
PubMed1989

Marker profile of mesothelial cells versus ovarian carcinoma cells.

We investigated the marker profile of human ascitic and cultured mesothelial cells, and compared it to that of ovarian carcinoma cells which are related in terms of their histogenesis, unrelated colon carcinomas being used as reference. Mesothelial and ovarian carcinoma cells could not be distinguished by (intermediate) filament typing, using monoclonal antibodies (MAbs) to keratins, vimentins and desmins. Colon carcinomas differed from mesothelial cells and ovarian carcinomas by the absence of keratin-7 filaments. The epithelial marker BW 495/36 was completely negative on mesothelial cells and positive on all ovarian and colon carcinoma cells. While CEA was found on about 85% of all colon carcinomas, CEA expression on mesothelial cells and ovarian carcinoma cells was below 20%. The ovarian carcinoma markers (OV-TL 3, OV-TL 10, OC 125, MOV 18) were strongly positive on ovarian carcinomas and negative on colon carcinomas (or limited to traces of immunofluorescence on some samples). Although the mesothelial cells showed weak or negative reactivity with these markers, OC 125 antigen was found by immunoelectron microscopy on the surface of cultured mesothelial cells, and was shed in the culture supernatant at concentrations of 50, 28, and 25 CA 125 U/ml/10(4) positive cells. This suggests that mesothelial cells may be responsible for the synthesis of CA 125 in ascitic fluid. The data indicate that ovarian carcinomas, mesothelial cells and colon carcinomas can be distinguished using a combination of anti-keratin antibodies with BW 495/36 and anti-ovarian carcinoma markers.

Antibodies, Monoclonal↗
PubMed1989

Analysis of nonpolar insertion mutations in the trfA gene of IncP plasmid RK2 which affect its broad-host-range property.

Replication of broad-host-range plasmid RK2 requires the protein product(s) of the plasmid-encoded trfA gene to initiate replication at oriV, the vegetative replication origin. The trfA gene contains two translational starts which direct translation of two polypeptides, of 382 and 285 amino acids, which differ by the 97 amino acids at their N-terminus. Nonpolar insertions which abolish expression of the larger TrfA polypeptide but otherwise retain the trfA gene's normal expression signals severely reduce plasmid replication efficiency in Pseudomonas aeruginosa and to a lesser extent in Pseudomonas putida, but have very little effect in Escherichia coli. This indicates that the organization of the trfA gene, producing two polypeptides products, plays an important part in the broad-host-range of plasmid RK2 by providing a degree of flexibility in the way the plasmid's replication system interacts with host biochemistry.

Conjugation, Genetic↗
PubMed1989

Return of gonadotropic function in postpartum women during bromocriptine treatment.

The rapid postpartum return of gonadotropic function during bromocriptine treatment was investigated through hormonal measurements in 15 women. Luteinizing hormone (LH) was measured by a novel and specific assay that does not cross-react with human chorionic gonadotropin (hCG). Prolactin was effectively suppressed by bromocriptine and hCG was cleared within 2 weeks. During the first week, both follicle-stimulating hormone (FSH) and LH were undetectable. The return of FSH and LH secretion occurred in the second postpartum week and initially was characterized by a high FSH/LH ratio. The causes of the 1-week delay in resumption of pituitary gonadotropin secretion are not known, but endogenous opioids may play a role. The possible relation between the high FSH/LH ratio and low LH pulse frequency also is discussed.

Bromocriptine↗
PubMed1989

Characterization of a hormone-producing ovarian carcinoma cell line.

An ovarian carcinoma cell line (OTN 11) was produced from the ascitic fluid of a patient with a moderately to well differentiated papilliferous cystadenocarcinoma of the ovary. The cell line was characterized using electron microscopy karyotyping, immunohistochemical techniques with monoclonal antibodies against keratins as epithelial markers, and the monoclonal antibodies OV-TL 3 and OC 125 as ovarian carcinoma markers. These techniques revealed the epithelial and adenocarcinomatous nature of the cell line and the presence of ovarian carcinoma-related surface markers. The adenocarcinomatous nature of the cell line also became apparent after heterotransplantation of cell suspensions into nude mice and nude rats, in which adenomatous tumor structures were formed. These xenografts had the same ultrastructural and immunohistochemical properties as the cell line. Despite the adenocarcinomatous character of the tumor the cultured cells release estradiol into the culture medium. We may conclude that OTN 11 is an ovarian carcinoma cell line which has retained highly differentiated functions, such as the production of an ovarian hormone.

Adult↗
PubMed1989

Cloning and expression of Treponema pallidum antigens in Escherichia coli.

A library of Treponema pallidum genomic DNA fragments produced by partial Sau3A digestion was established in Escherichia coli K12 using the plasmid vector pAT153. The library was screened using immune syphilitic rabbit serum and six recombinant phenotypes expressing eight treponemal polypeptides were detected. With two exceptions, all the recombinant gene products were the same size as polypeptides detected on Western immunoblots of T. pallidum. The genes encoding three novel gene products, with molecular masses in SDS-PAGE of 42, 17 and 15.5 kDa, which had not been cloned previously from T. pallidum were also identified. Monoclonal antibodies which reacted with four of the eight recombinant polypeptides were generated.

Antibodies, Monoclonal↗
PubMed1989

Serum hormone levels in pregnant cynomolgus monkeys.

Serum levels of estradiol (E2), progesterone (P), prolactin (Prl), monkey chorionic gonadotropin (mCG), dehydroepiandrosterone sulfate (DHEAS), and cortisol (C) in pregnant cynomolgus monkeys are described. mCG, E2, and Prl patterns resembled those of pregnant rhesus monkeys. P patterns differed from data from the literature. DHEAS patterns did not follow E2 patterns. C levels did not change during pregnancy.

Animals↗
PubMed1988

Replication of mini RK2 plasmid in extracts of Escherichia coli requires plasmid-encoded protein TrfA and host-encoded proteins DnaA, B, G DNA gyrase and DNA polymerase III.

Soluble extracts of Escherichia coli capable of carrying out replication of the mini-RK2 derivative pCT461 have been prepared from cells carrying this plasmid or from plasmid-free bacteria. The latter are dependent upon exogenously added plasmid-encoded replication protein (TrfA) and require additional DnaA protein for optimum activity. This dependence upon DnaA was confirmed by the failure of DnaA-deficient cell extracts to support replication of pCT461 in the absence of added DnaA protein. Replication is unidirectional and begins at or near oriV, the vegetative replication origin of RK2. DNase I protection studies with purified TrfA indicate that this protein acts by binding to short (17 base-pairs) directly repeated DNA sequences present in oriV. The in vitro replication is resistant to rifampicin but can be abolished by antibodies against DnaG protein (E. coli primase) or DnaB protein (helicase) and by DNA gyrase inhibitors. Inhibition by arabinosyl-CTP suggests that DNA polymerase III is responsible for elongation of nascent DNA strands. These results are discussed in relation to the mechanism of RK2 replication and in the context of the host range of the plasmid.

Bacterial Proteins↗
PubMed1988

[The Discretest].

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Adolescent↗