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At least 109 records · Page 6Linked to original sources
PubMed1990

Structures of the asparagine-linked oligosaccharides of an alkaline phosphatase, kasahara isozyme, purified from FL amnion cells.

Asparagine-linked oligosaccharides were quantitatively released by hydrazinolysis from an alkaline phosphatase, Kasahara isozyme, which was purified from FL amnion cells. Almost all of the oligosaccharides (98%) were acidic components, all of which can be converted to neutral oligosaccharides upon sialidase digestion. Structural analysis of the oligosaccharides by sequential exoglycosidase digestion in combination with methylation analysis revealed that the alkaline phosphatase of FL cells contains sialylated mono-, bi-, tri-, and tetraantennary complex type sugar chains with the Gal beta 1----4GlcNAc beta 1---- outer chains. Some of the tetraantennary sugar chains contain a single Gal beta 1----4GlcNAc beta 1----3Gal beta 1----4GlcNAc beta 1---- outer chain on their Man alpha 1----6 arm. Both fucosylated and nonfucosylated trimannosyl cores were found in the sugar chains. However, it is of interest that the core portion of monoantennary oligosaccharide was not fucosylated and that of the tetraantennary oligosaccharide with a tetrasaccharide outer chain was completely fucosylated.

Alkaline Phosphatase↗
PubMed1985

A simple method for staining of nucleoproteide equivalents in human FL cell chromosomes.

Chick embryo cell monolayers (CEC) treated by acidified thiazine dyes (pH 5 to 3) arrived into a mitogenic state. The diffuse dispersed chromatin of the nuclei clustered on the nuclear membranes and on the nucleoli in chromosome-like figures is usually estimated by electron microscopy. The clustering process could better be followed by light microscopy. The same treatment with acidified thiazine dyes on Fogh and Lund (FL) cell monolayers led unequivocally to induction of pre-formed chromosomes in interphasic cells with the same location as in the CEC. On the other hand, in metaphasic nuclei of FL cells, the above stain showed a cluster of deeply stained granules and pale stained strings. The last cluster model could be observed in the interior of metaphasic chromosomes aligned along the chromosomal arms in longitudinal, horizontal and/or crisscross position. These cluster patterns differ from conventional chromosomal bands obtained by Giemsa of fluorescent staining.

Animals↗
PubMed1980

Detection of type D retrovirus in a human amnion cell line (FL).

This paper describes the detection of viral particles in a human amnion cell line (FL). These particles belong to the group of type D retroviruses, because of their characteristic morphology, their major antigenic determinants, and the presence of particle-associated Mg++-preferring RNA-dependent DNA polymerase. In addition to budding and mature type D virus particles, intracytoplasmic type A structures have also been found. Immunological analysis provided evidence of cross-reactivity between the particles described and the Mason-Pfizer monkey virus as well as type D retroviruses from other human cell lines (PMF, HEp-2). Particles isolated from FL cell medium were shown to infect type D virus-susceptible human TU 197 cells.

Amnion↗
PubMed1978

[Karyologic study of transplantable cell lines. II. Comparative analysis of lines HEp-2, FL and RH].

A comparative karyologic analysis of three continuous cell lines of different origin (HEp-2, FL and RH) has shown them differing from each other both in the number of chromosomes, and in marker chromosomes. In cultures HEp-2, the modal class consists of cells possessing 58-59 chromosomes, cells in cultures Fl and RH have 60 and 66 chromosomes, resp. Marker chromosomes both similar and not similar, i.e. characteristic of a given cell line only were found in all the investigated cell lines. The data obtained do not exclude a possibility of contamination of studied continuous cell cultires with HeLa cell line.

Amnion↗
PubMed1981

Inhibition of the formation of electrical cell coupling of FL cell by tumor promoters.

The effect of tumor promoters on the formation of electrical cell coupling between human FL cells was investigated by means of a microelectrode technique. In culture medium containing 100 ng/ml of 12-O-tetradecanoyl phorbol-13-acetate (TPA), only 6% of the cells were coupled after 24 hr while more than 90% of the cells were coupled in control medium. The formation of cell coupling was also inhibited by other tumor promoters, phorbol-12,13-didecanoate, ingenol-13, 20-dibenzoate and mezerein, whereas non-promoting derivatives, phorbol and 4 alpha-phorbol-12, 13 didecanoate did not affect cell coupling formation. 7, 12-Dimethylbenz [a] anthracene, benzo [a] pyrene and N-methyl-N'-nitro-N-nitrosoguanidine did not affect the cell coupling. None of the chemicals used changed any electrical properties of the cell membrane, such as membrane potential and membrane resistance, of FL cells. Thus, it appears that TPA and other related tumor promoters inhibit the formation of cell coupling without changing the physiological properties of the membrane significantly.

Amnion↗
PubMed2004

Proteomic analysis of cellular responses to low concentration N-methyl-N'-nitro-N-nitrosoguanidine in human amnion FL cells.

We have shown previously that exposure to a low concentration of N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) induces comprehensive changes in the protein expression profile of human amnion FL cells, including the induction, suppression, upregulation, and downregulation of various proteins. In addition, by proteomic analysis combining two-dimensional gel electrophoresis (2-DE) and mass spectrometry, some of the induced and suppressed proteins were identified. In this study, we identified an additional 18 proteins among those that were either up- or downregulated by MNNG treatment. The proteins identified were a heterogeneous group that included several zinc finger proteins, proteins involved in signal transduction, cytoskeletal proteins, cell-cycle regulation proteins, and proteins with unknown functions. The involvement of these proteins in the cellular responses to alkylating agents has not been reported before and their physiological relevance is not clear. Therefore, our findings may help better understand the global cellular stress responses to chemical carcinogens, and may lead to new studies on the functions of these MNNG-responsive proteins. Furthermore, some of these proteins may serve as biomarkers for detecting exposure of human populations to environmental carcinogens.

Amnion↗
PubMed1982

Defective expression of mouse adenovirus Fl in human cells.

HeLa cells infected with mouse adenovirus strain Fl ( AdFl ) produce at least 2000 times less virus than permissive mouse 3T3 cells. Viral DNA synthesis, however, proceeds unimpaired. The defect in virion production was linked to a dramatic reduction in the synthesis of AdFl structural proteins, in particular the hexon. The identity of the AdFl hexon gene product was recognized through its immunogenic reactivity towards an antiserum raised against the human adenovirus type 2 (Ad2) hexon gene product. This cross-reactivity is reflected in an extensive DNA sequence homology between AdFl and Ad2 DNA at position 53-60, the locus of the hexon gene, on the Ad2 physical map. Through hybridization at different formamide concentrations, the present study identifies one additional, highly conserved region at map positions 14-15 on the Ad2 genome.

Adenoviridae↗
PubMed1993

Evidence for uptake2-mediated O-methylation of noradrenaline in the human amnion FL cell-line.

The uptake and metabolism of 3H-noradrenaline has been examined in the FL cell-line derived originally from human amnion. Cell cultures metabolised 3H-noradrenaline (1.0 mumol/l) to 3H-normetanephrine and, to a lesser extent, to metabolites (not distinguished) of the O-methylated deaminated fraction; primary deaminated metabolites were not detected. 3H-normetanephrine formation a) was not saturable in the noradrenaline concentration range 0.2-150 mumol/l, b) was decreased to 20%-30% of control levels by uptake2 inhibitors (O-methylisoprenaline, 20 and 100 mumol/l; cimetidine, 10 mumol/l; hydrocortisone, 200 mumol/l) and c), was almost insensitive to uptake1 inhibitors (cocaine, 30 mumol/l; desipramine, 3 mumol/l). Uptake of noradrenaline was manifested after 30 minutes as a 6-fold increase in the cell content of the amine following inhibition of catechol-O-methyl transferase, either alone or in conjunction with inhibition of monoamine oxidase. Uptake was decreased maximally to 40% of control levels by O-methylisoprenaline. IC50 values for inhibition of the O-methylisoprenaline-sensitive component of uptake were (in mumol/l): corticosterone (0.3), papaverine (1.1), O-methylisoprenaline (3.0), cimetidine (6.0), (-)noradrenaline (460), and tetraethylammonium (2230). Except for the last agent, for which a comparative value is not available, the IC50's are in good agreement with those for inhibition of uptake2 in the Caki-1 cell-line reported by other investigators. The component of uptake resistant to O-methylisoprenaline was depressed by papaverine (a 50% decrease at 50 mumol/l), but was not affected by the other uptake2 inhibitors or by cocaine (30 mumol/l).(ABSTRACT TRUNCATED AT 250 WORDS)

Amnion↗
PubMed1977

Biological and biophysical characteristics of mouse adenovirus, strain FL.

Mouse adenovirus, strain FL (MAV), is best propagated in primary rather than secondary cultures of mouse kidney cells. Virus release from the cells into the medium is fairly efficient; the average yield per cell is about 1000 TCID50. Morphological and other biophysical characteristics are those of a typical adenovirus: the icosahedral shell with a diameter of 74 nm, fiber projections of 29 nm length, its localization inside the infected kidney cells, a buoyant density in CsC1 of 1.34 g/ml, inhibition of multiplication by inhibitors of DNA synthesis. The thermostability is higher than that of human adenoviruses, whereas, in contrast to these, MAV is inactivated to a great extent by trypsin. Complete or incomplete hemagglutinin or toxin-like activity were not detected. The virus shows no cross-neutralization with human adenoviruses and a one-side cross-reaction with another murine adenovirus strain K-87. Soluble complement-fixing antigen of MAX exhibits a sedimentation rate of 12S identical to the hexon component of human adenoviruses; both antigens show a partial antigenic relationship in tests with appropriate antisera.

Adenoviridae↗
PubMed2000

Supercritical fluid extraction of polycyclic aromatic hydrocarbons from liver samples and determination by HPLC-FL.

An extraction/clean-up procedure by SFE was developed for isolating PAHs from liver samples for subsequent HPLC-FL determination of ten PAHs in the enriched extract. Recoveries (90-115%) and RSD % (< or =7.7) were satisfactory. When applied to 11 samples of bird of prey (Tyto alba) protected species and classified of special interest, from the Galicia (Northwest to Spain), benzo[ghi]perylene and indeno[1,2,3-cd]pyrene were undetectable; chrysene and benzo[a]pyrene are only detected in one sample; benzo[a]anthracene and benzo[k]fluoranthene are only quantified in one sample and benzo[b]fluoranthene in two samples. The other PAHs, anthracene, fluoranthene and pyrene are present in almost all the samples.

Animals↗
PubMed1992

Escherichia coli H(+)-ATPase: role of the delta subunit in binding Fl to the Fo sector.

The roles of the Escherichia coli H(+)-ATPase (FoFl) delta subunit (177 amino acid residues) was studied by analyzing mutants. The membranes of nonsense (Gln-23----end, Gln-29----end, Gln-74----end) and missense (Gly-150----Asp) mutants had very low ATPase activities, indicating that the delta subunit is essential for the binding of the Fl portion to Fo. The Gln-176----end mutant had essentially the same membrane-bound activity as the wild type, whereas in the Val-174----end mutant most of the ATPase activity was in the cytoplasm. Thus Val-174 (and possibly Leu-175 also) was essential for maintaining the structure of the subunit, whereas the two carboxyl terminal residues Gln-176 and Ser-177 were dispensable. Substitutions were introduced at various residues (Thr-11, Glu-26, Asp-30, Glu-42, Glu-82, Arg-85, Asp-144, Arg-154, Asp-161, Ser-163), including apparently conserved hydrophilic ones. The resulting mutants had essentially the same phenotypes as the wild type, indicating that these residues do not have any significant functional role(s). Analysis of mutations (Gly-150----Asp, Pro, or Ala) indicated that Gly-150 itself was not essential, but that the mutations might affect the structure of the subunit. These results suggest that the overall structure of the delta subunit is necessary, but that individual residues may not have strict functional roles.

Amino Acid Sequence↗
PubMed1983

Characterization of an epitope (determinant) structure in a developmentally regulated glycolipid antigen defined by a cold agglutinin Fl, recognition of alpha-sialosyl and alpha-L-fucosyl groups in a branched structure.

The antibody Fl shows preferential reactivity with adult erythrocytes over newborn erythrocytes, and its reactivity is abolished by sialidase treatment of the erythrocyte. The antibody was found to recognize binary determinants linked to the branched lacto-N-isooctaosylceramide (formula; see text) The presence of an N-acetylneuraminyl group at one end and L-fucosyl group at the other end is essential for the reactivity of the antibody. A substitution at the penultimate D-galactosyl residue of one of the chains with an alpha-D-(1 leads to 3)-linked 2-acetamido-2-deoxygalactosyl or galactosyl group did not inhibit the reactivity of the antibody. The new blood group A- and B-active, branched gangliosides are also isolated and characterized.

ABO Blood-Group System↗
PubMed1975

Possible identity between the hepatoma alkaline phosphatase and an isozyme of human amniotic membrane (FL cells).

The electrophoretically faster migrating alkaline phosphatase isozyme in FL cells derived from the human amniotic membrane was similar in some of the enzymatic and immunochemical properties to the variant alkaline phosphatase which occurs in serum and cancerous tissue of some patients with hepatocellular carcinoma. These facts suggest that the hepatoma-associated alkaline phosphatase may be the resurgence of the isozyme originally present in the amniotic membrane.

Alkaline Phosphatase↗
PubMed2005

FL-CTL assay: fluorolysometric determination of cell-mediated cytotoxicity using green fluorescent protein and red fluorescent protein expressing target cells.

Cytotoxic T lymphocytes (CTLs) are crucial effectors against intracellular pathogens and cancer. Accurate and efficient assessment of CTL activity is important for basic and clinical studies. Widely used CTL assays, including the chromium release, JAM test and ELISPOT, involve either radioisotopes or lengthy procedures. Here, we developed a new fluorolysometric CTL assay based on cell-mediated cytolysis of fluorescent protein (GFP or DsRed) expressing cells quantified by one of the fluoro-based methods: flow cytometry, fluorescence microplate reader, or fluorescence microscopy. With flexible detection methods and lentiviral vector transduced stable lines of either GFP+ or DsRed+ cells as targets for antigen presentation and equal number of the other as internal reference for consistency and accuracy, this assay is easy to perform and to scale-up for simultaneous multi-sample analyses. Using two different antigen systems, we demonstrated that this assay is very sensitive to determine primary CTL activity of both in vitro and in vivo primed antigen-specific T cells. Thus, this FL-CTL assay is highly sensitive, reliable, reproducible, economical, convenience and supports broad applications compared to conventional CTL assays.

Animals↗
PubMed1995

Raman spectroscopy of the filamentous virus Ff (fd, fl, M13): structural interpretation for coat protein aromatics.

Site-specific isotope substitutions in the coat protein (pVIII) of the filamentous bacterial virus Ff (fd, fl, M13) have been employed to advance vibrational band assignments and facilitate structural interpretation of the Raman spectrum. We report spectra of phage fd assembled in vivo from pVIII subunits incorporating either deuteriophenylalanine (Fd5), deuteriotryptophan (Wd5), or deuteriotyrosine (Yd4) residues with labeled ring sites. The deuterated aromatics were introduced into fd individually and in combination. On the basis of observed isotope shifts, definitive assignments have been developed for all prominent Raman bands diagnostic of the pVIII aromatic residues (F11, F42, F45, W26, Y21, Y24). The present study constitutes the first direct experimental determination of Raman fingerprints of tyrosine and phenylalanine side chains within hydrophobic alpha-helical domains and yields unexpected results. Importantly, neither Y21 nor Y24 of pVIII exhibits the "canonical" Fermi doublet expected in the 820-860 cm-1 interval of the Raman spectrum. Instead, each tyrosine exhibits a single band near 853 cm-1. Since the application of denaturing conditions is sufficient to generate in fd an apparent Fermi doublet, it is concluded that the anomalous singlet is intrinsic to tyrosine environments in the native virion assembly. In addition, the Raman results clearly demonstrate an interdependence of the environments of aromatic side chains in virion subunits. We show that the results on fd isotopomers are also confirmed by Raman spectroscopy of Ff virions incorporating the tyrosine mutations Y21M, Y24M, and Y21F/Y24S. The Raman marker bands identified for pVIII aromatics modify and extend Raman correlations proposed previously for proteins. The unusual environments detected for aromatic residues in the mature Ff assembly are discussed in relation to recently proposed models for filamentous virion architecture.

Amino Acid Sequence↗
PubMed2003

Assays for hydrophilic and lipophilic antioxidant capacity (oxygen radical absorbance capacity (ORAC(FL))) of plasma and other biological and food samples.

Methods are described for the extraction and analysis of hydrophilic and lipophilic antioxidants, using modifications of the oxygen radical absorbing capacity (ORAC(FL)) procedure. These methods provide, for the first time, the ability to obtain a measure of "total antioxidant capacity" in the protein free plasma, using the same peroxyl radical generator for both lipophilic and hydrophilic antioxidants. Separation of the lipophilic and hydrophilic antioxidant fractions from plasma was accomplished by extracting with hexane after adding water and ethanol to the plasma (hexane/plasma/ethanol/water, 4:1:2:1, v/v). Lipophilic and hydrophilic antioxidants were efficiently partitioned between hexane and aqueous solvents. Conditions for controlling temperature effects and decreasing assay variability using fluorescein as the fluorescent probe were validated in different laboratories. Incubation (37 degrees C for at least 30 min) of the buffer to which AAPH was dissolved was critical in decreasing assay variability. Lipophilic antioxidants represented 33.1 +/- 1.5 and 38.2 +/- 1.9% of the total antioxidant capacity of the protein free plasma in two independent studies of 6 and 10 subjects, respectively. Methods are described for application of the assay techniques to other types of biological and food samples.

Adult↗
PubMed2006

Design and synthesis of a photocleavable fluorescent nucleotide 3'-O-allyl-dGTP-PC-Bodipy-FL-510 as a reversible terminator for DNA sequencing by synthesis.

DNA sequencing by synthesis (SBS) using reversible fluorescent nucleotide terminators is potentially an efficient approach to address the limitations of current DNA sequencing techniques. Here, we report the design and synthesis of a 3'-O-allyl photocleavable fluorescent nucleotide analogue, 3'-O-allyl-dGTP-PC-Bodipy-FL-510, as a reversible terminator for SBS. The nucleotide is efficiently incorporated by DNA polymerase into a growing DNA strand to terminate the polymerase reaction. After that, the fluorophore is photocleaved quantitatively by irradiation at 355 nm, and the allyl group is rapidly and efficiently removed by using a Pd-catalyzed reaction under DNA-compatible conditions to regenerate a free 3'-OH group to reinitiate the polymerase reaction. Two cycles of such steps were successfully demonstrated to sequence a homopolymeric region of a DNA template, facilitating the development of SBS as a viable approach for high-throughput DNA sequencing.

Base Sequence↗