Detection of rare antigen-presenting cells by the lacZ T-cell activation assay suggests an expression cloning strategy for T-cell antigens.
AuthorsKarttunen J, Sanderson S, Shastri N
JournalProc Natl Acad Sci U S A
PubMed ID1378619
'The alpha/beta T-cell receptor a complex ligand formed by the association of antigenic peptides with molecules of the major histocompatibility complex (MHC). The inherent limitations of the conventional T-cell activation assays used to detect these peptide/MHC ligands have, until now, hampered the development of expression cloning systems for T-cell antigens. ... More
Primitive adult hematopoietic stem cells can function as osteoblast precursors.
AuthorsOlmsted-Davis EA, Gugala Z, Camargo F, Gannon FH, Jackson K, Kienstra KA, Shine HD, Lindsey RW, Hirschi KK, Goodell MA, Brenner MK, Davis AR
JournalProc Natl Acad Sci U S A
PubMed ID14673088
'Osteoblasts are continually recruited from stem cell pools to maintain bone. Although their immediate precursor is a plastic-adherent mesenchymal stem cell able to generate tissues other than bone, increasing evidence suggests the existence of a more primitive cell that can differentiate to both hematopoietic and mesenchymal cells. We show here ... More
Retroviral gene transfer is inhibited by chondroitin sulfate proteoglycans/glycosaminoglycans in malignant pleural effusions.
'Gene therapy may be an important adjuvant for treating cancer in the pleural space. The initial results of retroviral gene transfer to cancer cells in malignant pleural effusions revealed that transduction was markedly inhibited, and studies to characterize the inhibitory factor(s) were performed. The inhibition was contained within the soluble, ... More
In situ detection of transcriptionally active chromatin and genetic regulatory elements in individual viable mammalian cells.
AuthorsKerr WG, Nolan GP, Herzenberg LA
JournalImmunol Suppl
PubMed ID2807403
'Using a newly developed FACS method for quantifying the expression of the Escherischia coli lacZ reporter gene in viable mammalian cells, we have obtained cloned cell lines in which the expression of lacZ is under the control of native endogenous transcription elements. We infected the murine pre-B cell 70Z/3 with ... More
Transcriptional activity of the neuron-specific enolase (NSE) promoter in murine embryonic stem (ES) cells and preimplantation embryos.
'Mouse embryonic stem (ES) cells were transfected with a plasmid composed of an E. coli lacZ gene fused to 1.8 kb of rat neuron-specific enolase (NSE) promoter sequences. While this reporter construct had been shown previously to function exclusively in postmitotic neurons and neuro-endocrine cells of transgenic mice, stably transfected ... More
Ion channels, Ca2+ signaling, and reporter gene expression in antigen-specific mouse T cells.
AuthorsKerschbaum HH, Negulescu PA, Cahalan MD
JournalJ Immunol
PubMed ID9257822
'Whole cell recordings were performed in parallel with measurements of intracellular Ca2+ ([Ca2+]i) and gene expression using the murine T cell hybridoma, B3Z, a cell line stably infected with a lacZ reporter gene, driven by the minimal IL-2 promoter (NF-AT, nuclear factor of activated T cells). The physiologic roles of ... More
Localized sampling of cytoplasm from Xenopus oocytes for capillary electrophoresis.
AuthorsLuzzi V, Lee CL, Allbritton NL
JournalAnal Chem
PubMed ID9406526
'Continued progress in cellular physiology requires new measurement strategies which can be applied to solitary cells. Since many cellular signaling pathways act on time scales of a few seconds, there is a critical need for single-cell techniques with subsecond time resolution. Capillary electrophoresis shows great promise as a tool for ... More
GFAP gene expression during development of astrocyte.
AuthorsBaba H, Nakahira K, Morita N, Tanaka F, Akita H, Ikenaka K
JournalDev Neurosci
PubMed ID9078433
'Glial fibrillary acidic protein (GFAP) is expressed exclusively in astrocytes in the central nervous system. In order to characterize individual cultured cells in which the GFAP promoter is active and to identify the regulatory mechanisms of GFAP expression in these cells, we have developed a unique assay system for promoter ... More
Structural elements in domain IV that influence biophysical and pharmacological properties of human alpha1A-containing high-voltage-activated calcium channels.
AuthorsHans M, Urrutia A, Deal C, Brust PF, Stauderman K, Ellis SB, Harpold MM, Johnson EC, Williams ME
JournalBiophys J
PubMed ID10049321
'We have cloned two splice variants of the human homolog of the alpha1A subunit of voltage-gated Ca2+ channels. The sequences of human alpha1A-1 and alpha1A-2 code for proteins of 2510 and 2662 amino acids, respectively. Human alpha1A-2alpha2bdeltabeta1b Ca2+ channels expressed in HEK293 cells activate rapidly (tau+10mV = 2.2 ms), deactivate ... More
Transcriptional properties of genomic transgene integration sites marked by electroporation or retroviral infection.
AuthorsBaer A, Schübeler D, Bode J
JournalBiochemistry
PubMed ID10852701
'As a possible consequence of their survival strategy, proviruses are predominantly found in transcription-promoting genomic sites. For certain applications, these findings have led to the preferential use of retroviral vectors for the stable integration of transgenes. This study demonstrates that transcription levels of single-copy proviruses, which have been established either ... More
Plasmodium falciparum: modulation of surface antigenic expression of infected erythrocytes as revealed by cell fluorescence ELISA.
AuthorsBaruch D, Glickstein H, Cabantchik ZI
JournalExp Parasitol
PubMed ID1959571
'The surface reactivity of heterologous immune sera with erythrocytes infected with Plasmodium falciparum has been difficult to assess in quantitative terms because of the restricted accessibility of surface epitopes and the lack of sensitive methodologies. In a previous study we showed that cryptic antigens can become exposed on the surface ... More
Abnormal regulation of low density lipoprotein-sensitive events in a cholesterol transport mutant.
AuthorsDahl NK, Gutheil WG, Liscum L
JournalJ Biol Chem
PubMed ID8349588
'We have isolated and characterized Chinese hamster ovary cell mutants defective in the intracellular transport of low density lipoprotein (LDL)-derived cholesterol (Dahl, N. K., Reed, K. L., Daunais, M. A., Faust, J. R., and Liscum, L. (1992) J. Biol. Chem. 267, 4889-4896). Mutant 2-2, which exhibits a cholesterol transport defect ... More
Miniaturization of a functional transcription assay in yeast (human progesterone receptor) in the 384- and 1536-well plate format.
AuthorsBerg M, Undisz K, Thiericke R, Moore T, Posten C
JournalJ Biomol Screen
PubMed ID10803606
'Miniaturization of high throughput screening assays to high-density microplate formats (384 or 1536 wells) is currently the focus of research activity in modern drug discovery facilities. In this article, we describe the adaptation of a fluorescence-based functional transcription assay in yeast for assessing modulators of human progesterone receptor to the ... More
Reliable transient promoter assay using fluorescein-di-beta-D-galactopyranoside substrate.
AuthorsIkenaka K, Fujino I, Morita N, Iwasaki Y, Miura M, Kagawa T, Nakahira K, Mikoshiba K
JournalDNA Cell Biol
PubMed ID1693513
'The promoter region of the mouse myelin proteolipid protein (PLP) gene was cloned into a promoter testing vector, pIP111. The pIP111 vector is a promoterless derivative of pCH110 (SV40 early region promoter-lacZ) and contains the Escherichia coli lpp transcription terminator sequence at the 5'' end of the cloning site. The ... More
Cell tracing shows the contribution of the yolk sac to adult haematopoiesis.
AuthorsSamokhvalov IM, Samokhvalova NI, Nishikawa S
JournalNature
PubMed ID17377529
'The first haematopoietic stem cells (HSCs) appear in the aorta-gonad-mesonephros (AGM) region, major vitelline and umbilical vessels, and placenta; however, whether they arise locally or from immigrant yolk sac precursor cells remains unclear. This issue is best addressed by measuring cell-lineage relationships rather than cell potentials. To undertake long-term in ... More
Kinetic fluorescence measurement of fluorescein di-beta-D-galactoside hydrolysis by beta-galactosidase: intermediate channeling in stepwise catalysis by a free single enzyme.
AuthorsHuang ZJ
JournalBiochemistry
PubMed ID1909566
'Kinetic fluorescence measurements were employed to quantitative to stepwise hydrolysis of fluorescein di-beta-D-galactoside (FDG) by beta-galactosidase and the intermediate fluorescein mono-beta-D-galactoside (FMG) channeling. The kinetic parameters, Michaelis-Menten constant Km and enzymatic catalysis rate k2, for FDG hydrolysis to FMG by beta-galactosidase were obtained as 18.0 microM and 1.9 mumol.(min-mg)-1, respectively. ... More
Activity of mycobacterial promoters during intracellular and extracellular growth.
'pUS933, a bifunctional Mycobacterium-Escherichia coli translational fusion vector containing an amino-terminally truncated E. coli lacZ reporter gene, was constructed. Derivatives of pUS933, containing the promoter, RBS and start codon of the Mycobacterium bovis BCG hsp60 gene, the Mycobacterium leprae 28 kDa gene and the M. leprae 18 kDa gene were ... More
Simultaneous detection of beta-galactosidase activity and surface antigen expression in viable haematopoietic cells.
AuthorsBerger CN, Tan SS, Sturm KS
JournalCytometry
PubMed ID7851157
'The quantitation of intracellular beta-galactosidase activity has been described for viable cells. By using the fluorogenic substrate fluorescein-di-beta-D-galactopyranoside (FDG) in conjunction with flow cytometry, the proportion of positive cells as well as the level of expression can be determined. In this paper we describe beta-galactosidase expression in lymphoid and myeloid ... More
Epidermal growth factor receptor dimerization monitored in live cells.
AuthorsBlakely BT, Rossi FM, Tillotson B, Palmer M, Estelles A, Blau HM
JournalNat Biotechnol
PubMed ID10657132
'We present a method for monitoring receptor dimerization at the membrane of live cells. Chimeric proteins containing the epidermal growth factor (EGF) receptor extracellular and transmembrane domains fused to weakly complementing beta-galactosidase (beta-gal) deletion mutants were expressed in cells in culture. Treatment of the cells with EGF-like compounds for as ... More
Use of digitized video microscopy with a fluorogenic enzyme substrate to demonstrate cell- and compartment-specific gene expression in Salmonella enteritidis and Bacillus subtilis.
'A rapid and sensitive method for detection of cell- and compartment-specific gene expression in individual cells of both Gram-negative and Gram-positive microorganisms is described. The method combines the use of gene fusions to lacZ, and a fluorogenic beta-galactosidase substrate, fluorescein-di-(beta-D-galactopyranoside), with digitized video microscopy. All of the reporter constructs tested ... More
Retrovirus-mediated insertion of expressed and non-expressed genes at identical chromosomal locations.
AuthorsBerwin B, Barklis E
JournalNucleic Acids Res
PubMed ID8506135
'During retrovirus replication, a cellularly derived tRNA is annealed to the viral RNA at the primer binding site (PBS) to prime reverse transcription, and both the tRNA and the PBS become copied and matched together on complementary proviral DNA strands prior to integration. Using a viral PBS single base pair ... More
Bacillus subtilis spoIIIE protein required for DNA segregation during asymmetric cell division.
AuthorsWu LJ, Errington J
JournalScience
PubMed ID8160014
'Sporulation in Bacillus subtilis begins with an asymmetric cell division, producing a smaller prespore and a larger mother cell, both of which contain intact copies of the chromosome. The spoIIIE gene is required for chromosome segregation into the prespore compartment. The effects of the spoIIIE36 mutation on sigma F-dependent transcription ... More
Enzyme-linked immunoassay. I. Novel method for synthesis of the insulin-beta-D-galactosidase conjugate and its applicability for insulin assay.
AuthorsKato K, Hamaguchi Y, Fukui H, Ishikawa E
JournalJ Biochem (Tokyo)
PubMed ID1104600
'Pork insulin was subjected to mercaptosuccinylation and then coupled to beta-D-galactosidase [EC 3.2.1.23] from Escherichia coli using N,N''-o-phenylenedimaleimide. The competitive binding of the conjugate and insulin to anti-insulin antibody was tested. Results showed that formation of an insulin-beta-D-galactosidase conjugate could be used for immunoassay of insulin.' ... More
Beta-galactosidase activity in bacteria measured by flow cytometry.
AuthorsAlvarez AM, Ibáñez M, Rotger R
JournalBiotechniques
PubMed ID8292348
Fluorescent probes for living cells.
AuthorsJohnson I
JournalHistochem J
PubMed ID10188922
'The functional characteristics of fluorescent probes used for imaging and measuring dynamic processes in living cells are reviewed. Initial consideration is given to general design requirements for delivery, targeting, detectability and fluorescence readout, and current technologies for attaining them. Discussion then proceeds to the more application-specific properties of intracellular ion ... More
Identification of novel pheromone-response regulators through systematic overexpression of 120 protein kinases in yeast.
AuthorsBurchett SA, Scott A, Errede B, Dohlman HG
JournalJ Biol Chem
PubMed ID11337509
'Protein kinases are well known to transmit and regulate signaling pathways. To identify additional regulators of the pheromone signaling apparatus in yeast, we evaluated an array of 120 likely protein kinases encoded by the yeast genome. Each kinase was fused to glutathione S-transferase, overexpressed, and tested for changes in pheromone ... More
The inter-relatedness and interdependence of mouse T cell receptor gammadelta+ and alphabeta+ cells.
AuthorsPennington DJ, Silva-Santos B, Shires J, Theodoridis E, Pollitt C, Wise EL, Tigelaar RE, Owen MJ, Hayday AC
JournalNat Immunol
PubMed ID14502287
'Although T cell receptor (TCR)gammadelta+ and TCRalphabeta+ cells are commonly viewed as functionally independent, their relatedness and potential interdependence remain enigmatic. Here we have identified a gene profile that distinguishes mouse gammadelta cell populations from conventional alphabeta T cells. However, this profile was also expressed by sets of unconventional alphabeta ... More
A rapid enzyme assay for beta-galactosidase using optically gated sample introduction on a microfabricated chip.
AuthorsXu H, Ewing AG
JournalAnal Bioanal Chem
PubMed ID14634707
'The ability to perform enzyme assays on microchips is demonstrated using optically gated sample introduction. The hydrolysis of fluorescein mono- beta- d-galactopyranoside (FMG) by beta- d-galactosidase ( beta-Gal) is continuously monitored using a microchip for 5 to 10 min. The outcome of the reaction was analyzed by performing serial on-chip ... More
Squalene synthase-deficient mutant of Chinese hamster ovary cells.
AuthorsBradfute DL, Silva CJ, Simoni RD
JournalJ Biol Chem
PubMed ID1526971
'Squalene synthase (farnesyldiphosphate:farnesyldiphosphate farnesyltransferase, EC 2.5.1.21) converts farnesyl pyrophosphate to squalene, the first metabolic step committed solely to the biosynthesis of sterols. Using a fluorescence-activated cell sorting technique designed to screen for cells defective in the regulated degradation of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase, we isolated a squalene synthase-deficient mutant ... More
Defective transcription of the IL-2 gene is associated with impaired expression of c-Fos, FosB, and JunB in anergic T helper 1 cells.
AuthorsMondino A, Whaley CD, DeSilva DR, Li W, Jenkins MK, Mueller DL
JournalJ Immunol
PubMed ID8757326
'Anergic CD4+ Th cells do not produce IL-2 when challenged with Ag-pulsed accessory cells because of a transcriptional defect. In this work, we report that these anergic T cells are defective in their ability to up-regulate protein binding and transactivation at two critical IL-2 DNA enhancer elements: NF-AT (nuclear factor ... More
Assembly of GABAA receptor subunits: role of the delta subunit.
AuthorsSaxena NC, Macdonald RL
JournalJ Neurosci
PubMed ID7525894
'GABAA receptor channels (GABARs) composed of different combinations of rat alpha 1, beta 1, gamma 2L, and delta subunits were expressed transiently in mouse fibroblast cells (L929 cells). Whole-cell recordings were obtained from transfected cells to determine which combinations of GABAR subunits formed functional receptor channels, and to compare the ... More
Whole animal cell sorting of Drosophila embryos.
AuthorsKrasnow MA, Cumberledge S, Manning G, Herzenberg LA, Nolan GP
JournalScience
PubMed ID1898782
'Use of primary culture cells has been limited by the inability to purify most types of cells, particularly cells from early developmental stages. In whole animal cell sorting (WACS), live cells derived from animals harboring a lacZ transgene are purified according to their level of beta-galactosidase expression with a fluorogenic ... More
lacZ expression in germline transgenic zebrafish can be detected in living embryos.
AuthorsLin S, Yang S, Hopkins N
JournalDev Biol
PubMed ID8293887
'Use of transgenic technology in zebrafish has been limited by the inability to efficiently express transgenes in early embryos of F1 and subsequent generations and to rapidly detect transgenic fish. We generated transgenic fish by injecting fertilized eggs with the Escherichia coli lacZ gene under the control of the Xenopus ... More
Selection of bacterial virulence genes that are specifically induced in host tissues.
AuthorsMahan MJ, Slauch JM, Mekalanos JJ
JournalScience
PubMed ID8430319
'A genetic system was devised that positively selects for bacterial genes that are specifically induced when bacteria infect their host. With the pathogen Salmonella typhimurium, the genes identified by this selection show a marked induction in bacteria recovered from mouse spleen. Mutations in all ivi (in vivo-induced) genes that were ... More
AuthorsLorincz M, Roederer M, Diwu Z, Herzenberg LA, Nolan GP
JournalCytometry
PubMed ID8866216
'We report the development of a new fluorescence-activated cell sorter (FACS)-based reporter gene system utilizing the enzymatic activity of the E. coli beta-glucuronidase (gus) gene. When loaded with the Gus substrate fluorescein-di-beta-D-glucuronide (FDGlcu), individual mammalian cells expressing and translating gus mRNA liberate sufficient levels of intracellular fluorescein for quantitative analysis ... More
Beta-galactosidase activity in single differentiating bacterial cells.
AuthorsRusso-Marie F, Roederer M, Sager B, Herzenberg LA, Kaiser D
JournalProc Natl Acad Sci U S A
PubMed ID8396263
'Myxococcus xanthus strains containing transcriptional fusions to lacZ were analyzed and fractionated by differences in their levels of beta-galactosidase expression. The fluorogenic substrate for beta-galactosidase, fluorescein di-beta-galactopyranoside, was introduced into M. xanthus cells during a rapid decrease in osmolarity of the medium followed by a return to isoosmolarity. Fluorescein, the ... More
Rapid identification and quantitation of cells infected by recombinant herpesvirus (pseudorabies virus) using a fluorescence-based beta-galactosidase assay and flow cytometry.
AuthorsSaalmüller A, Mettenleiter TC
JournalJ Virol Methods
PubMed ID8227283
'We recently described construction and use of a beta-galactosidase expression cassette in isolating recombinant pseudorabies virus (PrV) mutants (Mettenleiter and Rauh, 1990). We report here the identification and exact quantitation of cells infected by these mutants using an assay based on the reaction of intracellular beta-galactosidase expressed during infection by ... More
Improved FACS-Gal: flow cytometric analysis and sorting of viable eukaryotic cells expressing reporter gene constructs.
AuthorsFiering SN, Roederer M, Nolan GP, Micklem DR, Parks DR, Herzenberg LA
JournalCytometry
PubMed ID1905992
'The previously reported FACS-Gal assay (Nolan et al., Proc Natl Acad Sci USA 85:2603-2607, 1988) measures E. coli lacZ-encoded beta-galactosidase activity in individual viable eukaryotic cells for a variety of molecular and cellular biological applications. Enzyme activity is measured by flow cytometry, using a fluorogenic substrate, which is hydrolyzed and ... More
AuthorsReddy S, Rayburn H, von Melchner H, Ruley HE
JournalProc Natl Acad Sci U S A
PubMed ID1495960
'Murine embryonic stem (ES) cells were infected with a retrovirus promoter trap vector, and clones expressing lacZ fusion genes (LacZ+) were isolated by fluorescence-activated cell sorting (FACS). Of 12 fusion genes tested, 1 was repressed when ES cells were allowed to differentiate in vitro. Two of three lacZ fusion genes ... More
A microplate fluorimetric assay for transfection of the beta-galactosidase reporter gene.
AuthorsRakhmanova VA, MacDonald RC
JournalAnal Biochem
PubMed ID9514778
A microspectrofluorometric study of porphyrin-photosensitized single living cells--I. Membrane alterations.
Fluorogenic substrates for beta-D-galactosidases and phosphatases derived from fluorescein (3,6-dihydroxyfluoran) and its monomethylether.
AuthorsROTMAN B, ZDERIC JA, EDELSTEIN M,
JournalProc Natl Acad Sci U S A
PubMed ID13975398
Syntheses of several new biochemically useful fluorogenic substrates for beta-D-galactosidases and phosphatases are described.
beta-D-galactosidase activity in single yeast cells during cell cycle of Saccharomyces lactis.
AuthorsYashphe J, Halvorson HO
JournalScience
PubMed ID1257751
Single Saccharomyces lactis cells taken from a random population were assayed for beta-D-galactosidase activity under a microscope equipped for fluorogenic measurements. The cells were also photographed, and enzymatic activity was correlated to the size of cell buds. A perodic pattern of enzyme synthesis was found during the cell cycle. ... More
A kinetic study on the enzymatic hydrolysis of fluorescein diacetate and fluorescein-di-beta-D-galactopyranoside.
AuthorsHofmann J, Sernetz M
JournalAnal Biochem
PubMed ID6614449
The kinetics of the hydrolysis of fluoresceindiacetate and fluorescein-di-beta-D-galactopyranoside were investigated by thin-layer chromatography. The time course of the concentrations of substrate, monosubstituted intermediate, and product was simulated numerically. The mathematical model takes into account the competition of substrate and intermediate and the accumulation of the intermediate at the enzyme. ... More
A simple and sensitive enzyme-mediated assay of biotin.
AuthorsHuang Z, Haugland RP, Szalecka D, Haugland RP
JournalBiotechniques
PubMed ID1476723
Free biotin was quantitated by a competition by coating biotin-bovine serum albumin conjugate on a polystyrene microplate for binding to avidin-beta-galactosidase conjugate. The enzyme conjugate remaining on the plate surface as a result of the competition was detected by reaction with one of the following fluorogenic substrates, resorufin beta-D-galactoside and ... More
Fluorescence-based enzymatic assay by capillary electrophoresis laser-induced fluorescence detection for the determination of a few beta-galactosidase molecules.
AuthorsCraig D, Arriaga EA, Banks P, Zhang Y, Renborg A, Palcic MM, Dovichi NJ
JournalAnal Biochem
PubMed ID7785765
beta-Galactosidase can be assayed by monitoring the generation of the fluorescent products, fluorescein-mono-beta-D-galactopyranoside and fluorescein, when the fluorogenic substrate fluorescein-di-beta-D-galactopyranoside is used. We have used capillary electrophoresis with ultrasensitive laser-induced fluorescence detection to monitor the formation of the fluorescent products of off-column enzymatic reactions. By analyzing as little as 40 ... More
Use of fluorescein-di-beta-D-galactopyranoside (FDG) and C12-FDG as substrates for beta-galactosidase detection by flow cytometry in animal, bacterial, and yeast cells.
AuthorsPlovins A, Alvarez AM, Ibañez M, Molina M, Nombela C
JournalAppl Environ Microbiol
PubMed ID7811104
Fluorescein-di-beta-D-galactopyranoside (FDG) was found to be a useful substrate for beta-galactosidase detection by flow cytometry in gram-negative bacteria, since it entered viable cells and gave a fluorescence emission proportional to the enzymatic activity. C12-FDG, a more lipophilic derivative, gave a very poor signal because of the lack of penetration. On ... More
Measurement of transcriptional activity in pathogenic bacteria recovered directly from infected host tissue.
AuthorsSlauch JM, Mahan MJ, Mekalanos JJ
JournalBiotechniques
PubMed ID8024783
In order to understand the genetic regulation of bacterial genes whose products are important for pathogenesis, one needs to measure the expression of the genes during the infection process. We have devised a method to measure the transcriptional activity of such genes from bacteria recovered directly from infected host tissue. ... More
Prevention of atherosclerosis in apolipoprotein E-deficient mice by bone marrow transplantation.
AuthorsLinton MF, Atkinson JB, Fazio S
JournalScience
PubMed ID7863332
Apolipoprotein E (apoE) deficiency causes severe hyperlipidemia and atherosclerosis in humans and in gene-targeted mice. Although the majority of apoE in plasma is of hepatic origin, apoE is synthesized by a variety of cell types, including macrophages. Because macrophages derive from hematopoietic cells, bone marrow transplantation was used to examine ... More
Increased atherosclerosis in mice reconstituted with apolipoprotein E null macrophages.
Macrophage-derived foam cells express apolipoprotein E (apoE) abundantly in atherosclerotic lesions. To examine the physiologic role of apoE secretion by the macrophage in atherogenesis, bone marrow transplantation was used to reconstitute C57BL/6 mice with macrophages that were either null or wild type for the apoE gene. After 13 weeks on ... More
Region-specific differentiation of the hippocampal stem cell line HiB5 upon implantation into the developing mammalian brain.
AuthorsRenfranz PJ, Cunningham MG, McKay RD
JournalCell
PubMed ID1878969
Proliferating precursors to the distinct cell types constituting the mammalian brain can be identified by the presence of the nestin intermediate filament. We report the establishment of a nestin-positive cell line, HiB5, from embryonic precursor cells to the rat hippocampus. Since it was immortalized using the temperature-sensitive allele tsA58 of ... More
Detection of acid-beta-galactosidase activity in viable human fibroblasts by flow cytometry.
AuthorsJongkind JF, Verkerk A, Sernetz M
JournalCytometry
PubMed ID3093179
The fluorogenic substrate fluorescein-di-beta-D-galactopyranoside was used to detect acid beta-galactosidase in intact cultured human fibroblasts. The accumulation of intracellular fluorescein, as measured by flow cytophotometry was linear with the incubation time in three control strains. The two fibroblast strains from patients with acid beta-galactosidase deficiency did not show an accumulation ... More
Targeted ablation of diverse cell classes in the nervous system in vivo.
AuthorsNirenberg S, Cepko C
JournalJ Neurosci
PubMed ID8340805
The study of both the function and development of complex neural systems would be greatly facilitated by a means for systematically blocking intercell communication. One way of preventing cells from signaling each other is to remove them from the system by ablation. Here we present a general technique for visualizing ... More
Isolation of mutant T lymphocytes with defects in capacitative calcium entry.
AuthorsSerafini AT, Lewis RS, Clipstone NA, Bram RJ, Fanger C, Fiering S, Herzenberg LA, Crabtree GR
JournalImmunity
PubMed ID7648396
Calcium and calcium-binding proteins play important roles in the signaling cascade leading from the initial engagement of TCRs on T cells to the fully activated state. To undertake a molecular dissection of this cascade, we first isolated a Jurkat T cell line derivative containing the NF-AT promoter element driving transcription ... More
Use of green fluorescent protein/Flp recombinase fusion protein and flow cytometric sorting to enrich for cells undergoing Flp-mediated recombination.
AuthorsSabath DE, Shim MH
JournalBiotechniques
PubMed ID10818704
Flp recombinase has been used extensively for in vivo manipulation of eukaryotic DNA at specific sequences designated as FRT sites. We developed a method to use Flp-mediated recombination without the need for drug resistance or metabolic selection of cells in which recombination has occurred. We generated expression plasmids directing expression ... More
Retroviral mediated gene transfer into bone marrow progenitor cells: use of beta-galactosidase as a selectable marker.
AuthorsStrair RK, Towle M, Smith BR
JournalNucleic Acids Res
PubMed ID2118622
Recombinant retroviruses have been utilized as vectors for gene transfer in model systems of gene therapy. Since many of these model systems require the transplantation of genetically modified primary cells it is important to devise methods which will allow the rapid and efficient selection for transplantation of only the cells ... More
Introduction of a lacZ reporter gene into the mouse int-2 locus by homologous recombination.
AuthorsMansour SL, Thomas KR, Deng CX, Capecchi MR
JournalProc Natl Acad Sci U S A
PubMed ID2120706
We demonstrate that the frequency of gene targeting is unaffected by the length of nonhomologous DNA transferred to a target chromosomal sequence. A result of this finding is that a much wider spectrum of designed genomic alterations is now feasible. As a first application, we inserted a 5.4-kilobase cassette of ... More
Microfabricated arrays of femtoliter chambers allow single molecule enzymology.
AuthorsRondelez Y, Tresset G, Tabata KV, Arata H, Fujita H, Takeuchi S, Noji H
JournalNat Biotechnol
PubMed ID15723045
Precise understanding of biological functions requires tools comparable in size to the basic components of life. Single molecule studies have revealed molecular behaviors usually hidden in the ensemble- and time-averaging of bulk experiments. Although most such approaches rely on sophisticated optical strategies to limit the detection volume, another attractive approach ... More
Retinoic acid is required early during adult neurogenesis in the dentate gyrus.
AuthorsJacobs S, Lie DC, DeCicco KL, Shi Y, DeLuca LM, Gage FH, Evans RM
JournalProc Natl Acad Sci U S A
PubMed ID16505366
Retinoic acid (RA) is commonly used in vitro to differentiate stem cell populations including adult neural stem cells into neurons; however, the in vivo function of RA during adult neurogenesis remains largely unexplored. We found that depletion of RA in adult mice leads to significantly decreased neuronal differentiation within the ... More
Nuclear targeting peptide scaffolds for lipofection of nondividing mammalian cells.
AuthorsSubramanian A, Ranganathan P, Diamond SL
JournalNat Biotechnol
PubMed ID10471928
Lipofection of nondividing cells is inefficient because much of the transfected DNA is retained in endosomes, and that which escapes to the cytoplasm enters the nucleus at low rates. To improve the final rate-limiting step of nuclear import, we conjugated a nonclassical nuclear localization signal (NLS) containing the M9 sequence ... More
Optical imaging fiber-based live bacterial cell array biosensor.
AuthorsBiran I, Rissin DM, Ron EZ, Walt DR
JournalAnal Biochem
PubMed ID12672418
A live cell array biosensor was fabricated by immobilizing bacterial cells on the face of an optical imaging fiber containing a high-density array of microwells. Each microwell accommodates a single bacterium that was genetically engineered to respond to a specific analyte. A genetically modified Escherichia coli strain, containing the lacZ ... More
Early determination of a mouse somatosensory cortex marker.
AuthorsCohen-Tannoudji M, Babinet C, Wassef M
JournalNature
PubMed ID8133892
The mammalian neocortex is subdivided into functionally distinct areas differing in cytoarchitecture and connectivity. Areal specification is thought to occur late in development and to be controlled by extrinsic cues, particularly thalamic afferents. We have produced a transgenic mouse line in which beta-galactosidase expression in the neocortex is largely restricted ... More
Human mesenchymal stem cells in rodent whole-embryo culture are reprogrammed to contribute to kidney tissues.
AuthorsYokoo T, Ohashi T, Shen JS, Sakurai K, Miyazaki Y, Utsunomiya Y, Takahashi M, Terada Y, Eto Y, Kawamura T, Osumi N, Hosoya T
JournalProc Natl Acad Sci U S A
PubMed ID15728383
The use of stem cells has enabled the successful generation of simple organs. However, anatomically complicated organs such as the kidney have proven more refractory to stem-cell-based regenerative techniques. Given the limits of allogenic organ transplantation, an ultimate therapeutic solution is to establish self-organs from autologous stem cells and transplant ... More
Endocytosis of major histocompatibility complex class I molecules is induced by the HIV-1 Nef protein.
AuthorsSchwartz O, Maréchal V, Le Gall S, Lemonnier F, Heard JM
JournalNat Med
PubMed ID8612235
Like other pathogenic viruses, HIV-1 down-modulates surface expression of major histocompatibility complex class I (MHC-I) molecules in infected cells, thus impairing lysis by cytotoxic T lymphocytes. We have observed that this phenomenon depends on the expression of Nef. nef is an early gene of primate lentiviruses, which is necessary for ... More
Selective encapsulation of single cells and subcellular organelles into picoliter- and femtoliter-volume droplets.
AuthorsHe M, Edgar JS, Jeffries GD, Lorenz RM, Shelby JP, Chiu DT
JournalAnal Chem
PubMed ID15762555
This paper describes a method, which combines optical trapping and microfluidic-based droplet generation, for selectively and controllably encapsulating a single target cell or subcellular structure, such as a mitochondrion, into a picoliter- or femtoliter-volume aqueous droplet that is surrounded by an immiscible phase. Once the selected cell or organelle is ... More
Channel electrophoresis for kinetic assays.
AuthorsLiu YM, Sweedler JV
JournalAnal Chem
PubMed ID8694257
A rectangular channel electrophoresis system and a cylindrical sampling capillary combination allows chemical changes in nanoliter-volume samples to be monitored as a function of time. The electrophoretic microseparation is carried out in a rectangular channel with a 7 -cm-long, 40-microm x 2.5-cm geometry and is coupled to a 50-microm-i.d. cylindrical ... More
Nucleic acid detection using non-radioactive labelling methods.
AuthorsMansfield ES, Worley JM, McKenzie SE, Surrey S, Rappaport E, Fortina P
JournalMol Cell Probes
PubMed ID7477006
Nucleic acid probe-based assays are now widely used in genetic research, human identification, forensics and in a broad spectrum of clinical assays in the fields of microbiology, haematology/oncology and virology. Labelled probes are used in a variety of assay formats including dot-blots, Southern blots (DNA target), Northern blots (RNA target), ... More
Retrovirally introduced antisense integrin RNA inhibits neuroblast migration in vivo.
AuthorsGalileo DS, Majors J, Horwitz AF, Sanes JR
JournalNeuron
PubMed ID1463609
We used retrovirus-mediated gene transfer to ask whether integrins are involved in the development of neuroblasts in the chicken optic tectum. Vectors were constructed with the E. coli lacZ gene in the sense orientation and beta 1 integrin sequences in the antisense orientation. Tests in culture showed that the progeny ... More
Verapamil inhibition of enzymatic product efflux leads to improved detection of beta-galactosidase activity in lacZ-transfected cells.
AuthorsPoot M, Arttamangkul S
JournalCytometry
PubMed ID9136753
The beta-galactosidase activity encoded by the lacZ gene of Escherichia coli is widely used to monitor successful expression of transfected genes. Fluorogenic substrates allow detection of enzyme activity in viable cells, which, subsequently, can be selected for further study on the basis of fluorescence emission. We analyzed three fluorogenic substrates ... More
Visualizing interleukin 2 gene expression at the single cell level.
AuthorsEmilie D, Peuchmaur M, Barad M, Jouin H, Maillot MC, Couez D, Nicolas JF, Malissen B
JournalEur J Immunol
PubMed ID2792182
To analyze the expression of the interleukin (IL)2 gene at the single cell level, we have constructed a chimeric gene in which the regulatory sequences from the mouse IL2 gene were fused 5' proximal to the coding region of the Escherichia coli beta-galactosidase (lacZ) gene. Once stably introduced into a ... More
Fluorescence-activated cell analysis and sorting of viable mammalian cells based on beta-D-galactosidase activity after transduction of Escherichia coli lacZ.
AuthorsNolan GP, Fiering S, Nicolas JF, Herzenberg LA
JournalProc Natl Acad Sci U S A
PubMed ID3128790
We demonstrate that individual cells infected with and expressing a recombinant retrovirus carrying the Escherichia coli beta-galactosidase gene (lacZ) can be viably stained, analyzed, sorted, and cloned by fluorescence-activated cell sorting based on the levels of lacZ expressed. To accomplish this we have devised a method to enzymatically generate and ... More
Flow cytometry sorting of viable bacteria and yeasts according to beta-galactosidase activity.
AuthorsNir R, Yisraeli Y, Lamed R, Sahar E
JournalAppl Environ Microbiol
PubMed ID2128011
We describe a novel method for quantitative measurement of beta-galactosidase (beta-gal) levels in bacteria and yeasts by using flow cytometry, a method which allows viable microbial cells to be sorted on the basis of the expressed activity and to be recultivated. The method is based on encapsulating single cells in ... More
Deoxyribonucleic acid vaccines encoding antigens with rapid proteasome-dependent degradation are highly efficient inducers of cytolytic T lymphocytes.
AuthorsWu Y, Kipps TJ
JournalJ Immunol
PubMed ID9550402
We generated plasmid expression vectors encoding ubiquitin and beta-galactosidase (beta-gal) with different intervening amino acids, allowing for the production of processed protein products that have either stabilizing or destabilizing residues at their N-termini. P815 cells transfected with plasmids encoding beta-gal with a destabilizing N-terminus did not have detectable expression beta-gal ... More
TNF-alpha pretreatment prevents subsequent activation of cultured brain cells with TNF-alpha and hypoxia via ceramide.
AuthorsGinis I, Schweizer U, Brenner M, Liu J, Azzam N, Spatz M, Hallenbeck JM
JournalAm J Physiol
PubMed ID10329967
We have developed a cellular model in which cultured astrocytes and brain capillary endothelial cells preconditioned with tumor necrosis factor-alpha (TNF-alpha) fail to upregulate intercellular adhesion molecule-1 (ICAM-1) protein (80% inhibition) and mRNA (30% inhibition) when challenged with TNF-alpha or exposed to hypoxia. Inasmuch as ceramide is known to mediate ... More
Flow cytometric study of differentiating cultures of Bacillus subtilis.
AuthorsChung JD, Conner S, Stephanopoulos G
JournalCytometry
PubMed ID7587720
We report on 1) the development of a flow cytometry-based technique for detecting beta-galactosidase in differentiating cultures of Bacillus subtilis and 2) the application of this technique in the study of early developmental gene expression. The problems associated with generating detectable signals (despite the small size of B. subtilis cells) ... More
Beta-galactosidase activity in transfected Ltk- cells is differentially regulated in monolayer and in spheroid cultures.
AuthorsKlünder I, Hülser DF
JournalExp Cell Res
PubMed ID8319768
We have investigated whether three-dimensional cultivation of cells to multicell spheroids influences the expression of a transfected gene. Ltk- cells (mouse fibroblasts, thymidine kinase negative) have been transfected with a bacterial lacZ gene which was coupled to a beta-actin promoter. The transfected cells synthesize beta-galactosidase, a cytoplasmic enzyme which can ... More
Characterization of the micro-environment of Salmonella typhimurium-containing vacuoles within MDCK epithelial cells.
Salmonella typhimurium has the capacity to enter into and multiply within epithelial cells. During the entire intracellular stage, bacteria are enclosed within a vacuole. To characterize the micro-environment of the bacteria-containing vacuoles, we have used a new method to measure the expression levels of several S. typhimurium genes in intracellular ... More
Visualization of CD4/CD8 T cell commitment.
AuthorsChan S, Correia-Neves M, Dierich A, Benoist C, Mathis D
JournalJ Exp Med
PubMed ID9858518
A system to innocuously visualize T cell lineage commitment is described. Using a "knock-in" approach, we have generated mice expressing a beta-galactosidase reporter in place of CD4; expression of beta-galactosidase in these animals appears to be an accurate and early indicator of CD4 gene transcription. We have exploited this knock-in ... More
Experiments in transgenic mice show that hepatocytes are the source for postnatal liver growth and do not stream.
AuthorsKennedy S, Rettinger S, Flye MW, Ponder KP
JournalHepatology
PubMed ID7541385
One hypothesis is that postnatal liver growth involves replication of mature hepatocytes, which have an unlimited proliferative potential. An alternative viewpoint is that only certain periportal cells can replicate extensively and that daughter cells stream slowly from the periportal to the pericentral region of the liver. Transgenic mice expressing the ... More
An enzyme-linked procedure for the detection and estimation of surface receptors on cells.
AuthorsCameron DJ, Erlanger BF
JournalJ Immunol
PubMed ID774980
An enzyme immunoassay procedure has been developed for the visualization and estimation of lymphocyte surface receptors. beta-Galactosidase (beta-gal'ase) was covalently linked to sheep anti-rabbit immunoglobulin (SARIg), to ovalbumin (OA), and to adenosine (A). Exposure of rabbit peripheral lymphocytes to SARIg-beta-gal'ase and subsequent incubation with the fluorogenic substrate fluorescein-beta-digalactopyranoside allowed visualization ... More
A fluorimetric method using fluorescein di-beta-D-galactopyranoside for quantifying the senescence-associated beta-galactosidase activity in human foreskin fibroblast Hs68 cells.
AuthorsYang NC, Hu ML
JournalAnal Biochem
PubMed ID14751269
The senescence-associated beta-galactosidase (SA-betaG) assay is one of the few accepted markers of cell aging. However, the cytochemical method using 5-bromo-4-chloro-3-indolyl beta-D-galactopyranoside (X-Gal) as substrate is limited in sensitivity and is only semiquantitative. Here, we modified the X-Gal method by replacing X-Gal with fluorescein di-beta-D-galactopyranoside (FDG) as substrate for SA-betaG, ... More
The adult CNS retains the potential to direct region-specific differentiation of a transplanted neuronal precursor cell line.
AuthorsShihabuddin LS, Hertz JA, Holets VR, Whittemore SR
JournalJ Neurosci
PubMed ID7472427
The chronic survival and differentiation of the conditionally immortalized neuronal cell line, RN33B, was examined following transplantation into the adult and neonatal rat hippocampus and cerebral cortex. In clonal culture, differentiated RN33B cells express p75NTR and trkB mRNA and protein, and respond to brain-derived neurotrophic factor treatment by inducing c-fos ... More
Construction and expression of an adenosine deaminase::lacZ fusion gene.
AuthorsShen Q, van Beusechem VW, Einerhand MP, Hendrikx PJ, Valerio D
JournalGene
PubMed ID1901822
A eukaryotic expression vector was constructed in which the coding nucleotide sequences (ADA) of human adenosine deaminase (ADA) were fused in frame with the coding sequences of the bacterial gene lacZ encoding beta-galactosidase (beta Gal). This ADA::lacZ fusion gene was anticipated to encode a hybrid protein that has retained the ... More
A microELISA assay for detection of anti-HLA activity of mouse monoclonal antibodies using an Astroscan 2100 automated plate reader.
A microenzyme-linked immunosorbent assay (microELISA) method has been developed using an Astroscan 2100 system automated plate reader which was initially designed for tissue typing by a two colour fluorescent microcytotoxicity assay. A 96-well plate ELISA used for screening mouse monoclonal antibodies raised against surface HLA antigens has been modified for ... More
Disruption of the adenosine deaminase (ADA) gene using a dicistronic promoterless construct: production of an ADA-deficient homozygote ES cell line.
AuthorsVaulont S, Daines S, Evans M
JournalTransgenic Res
PubMed ID7655514
In man, deficiency of ADA activity is associated with an autosomal recessive form of severe combined immunodeficiency (SCID), a disease with profound defects both cellular and humoral immunity. Current treatments of ADA deficient patients include bone marrow transplantation, enzyme replacement and somatic gene therapy. The mechanism of the selective immune ... More
Functional antagonism between Msx2 and CCAAT/enhancer-binding protein alpha in regulating the mouse amelogenin gene expression is mediated by protein-protein interaction.
AuthorsZhou YL, Lei Y, Snead ML
JournalJ Biol Chem
PubMed ID10859305
Ameloblast-specific amelogenin gene expression is spatiotemporally regulated during tooth development. In a previous study, the CCAAT/enhancer-binding protein alpha (C/EBPalpha) was identified as a transcriptional activator of the mouse amelogenin gene in a cell type-specific manner. Here, Msx2 is shown to repress the promoter activity of amelogenin-promoter reporter constructs independent of ... More
Characterization of T cell mutants with defects in capacitative calcium entry: genetic evidence for the physiological roles of CRAC channels.
AuthorsFanger CM, Hoth M, Crabtree GR, Lewis RS
JournalJ Cell Biol
PubMed ID7593187
Prolonged Ca2+ influx is an essential signal for the activation of T lymphocytes by antigen. This influx is thought to occur through highly selective Ca2+ release-activated Ca2+ (CRAC) channels that are activated by the depletion of intracellular Ca2+ stores. We have isolated mutants of the Jurkat human T cell line ... More
A transgenic mouse model for studying the lineage relationships and differentiation program of type II pneumocytes at various stages of lung development.
AuthorsHansbrough JR, Fine SM, Gordon JI
JournalJ Biol Chem
PubMed ID8486660
A pedigree of transgenic mice has been characterized that contains a H2-Kb/LacZ fusion gene that exhibits integration site-dependent expression from the earliest stages of lung development through adulthood. Histochemical and immunocytochemical studies indicate that the LacZ reporter appears throughout the pulmonary endoderm by embryonic day 11 (E11). A proximal-to-distal wave ... More
Assembly of GABAA receptor subunits: analysis of transient single-cell expression utilizing a fluorescent substrate/marker gene technique.
AuthorsAngelotti TP, Uhler MD, Macdonald RL
JournalJ Neurosci
PubMed ID7681869
GABAA receptor channels (GABARs) composed of varying combinations of alpha 1, beta 1, and gamma 2S subunits were transiently expressed in mammalian cell lines. The whole-cell patch-clamp recording technique was used to determine which combinations of GABAR subunits produced functional receptor channels and whether assembly of GABAR subunits into receptor ... More
Assembly of GABAA receptor subunits: alpha 1 beta 1 and alpha 1 beta 1 gamma 2S subunits produce unique ion channels with dissimilar single-channel properties.
AuthorsAngelotti TP, Macdonald RL
JournalJ Neurosci
PubMed ID7681870
Recent experimental evidence has led to the hypothesis that GABAA receptor channel (GABAR) heterogeneity or receptor channel subtypes may occur by differential assembly of a given set of subunits into various configurations. Alternatively, assembly of subunits into mature GABARs may arise from an ordered process to produce a preferred form ... More
Platelet-derived growth factor-induced alterations in vinculin distribution in porcine vascular smooth muscle cells.
AuthorsHerman B, Roe MW, Harris C, Wray B, Clemmons D
JournalCell Motil Cytoskeleton
PubMed ID3121190
Exposure of porcine vascular smooth muscle cells to platelet-derived growth factor (PDGF; 18-180 ng/ml) but not epidermal growth factor (EGF; 30 ng/ml), somatomedin C (SmC; 30 ng/ml), or insulin (10 microM), results in a rapid, reversible, time- and concentration-dependent disappearance of vinculin staining in adhesion plaques; actin-containing stress fibers also ... More
Primary mouse myoblast purification, characterization, and transplantation for cell-mediated gene therapy.
AuthorsRando TA, Blau HM
JournalJ Cell Biol
PubMed ID8207057
The transplantation of cultured myoblasts into mature skeletal muscle is the basis for a new therapeutic approach to muscle and non-muscle diseases: myoblast-mediated gene therapy. The success of myoblast transplantation for correction of intrinsic muscle defects depends on the fusion of implanted cells with host myofibers. Previous studies in mice ... More
No intermediate channelling in stepwise hydrolysis of fluorescein di-beta-D-galactoside by beta-galactosidase.
AuthorsFieldler F, Hinz H
JournalEur J Biochem
PubMed ID8200355
For the hydrolysis of the two glycosidic bonds of fluorescein di-beta-D-galactoside (FDG) by beta-galactosidase from Escherichia coli, small [Hofmann, J. & Sernetz, M. (1983) Anal. Biochem. 131, 180-186] to dramatic [Huang, Z. (1991) Biochemistry 30, 8535-8540] deviations from simple stepwise substrate-intermediate-product kinetics have been reported. Intermediate channelling, a preferred hydrolysis ... More
Intercellular signalling in Drosophila segment formation reconstructed in vitro.
AuthorsCumberledge S, Krasnow MA
JournalNature
PubMed ID8505983
Genetic studies show that intercellular signalling is involved in key steps in Drosophila melanogaster development, but it has not previously been possible to investigate these processes in simplified in vitro systems. Analysis of engrailed (en) and wingless (wg) and other segment polarity genes suggests that two or more intercellular signalling ... More
Measurement of ligand-induced activation in single viable T cells using the lacZ reporter gene.
AuthorsKarttunen J, Shastri N
JournalProc Natl Acad Sci U S A
PubMed ID1902576
We have used the bacterial beta-galactosidase gene (lacZ) as a reporter gene for the rapid measurement of T-cell antigen receptor (TCR)-mediated activation of individual T cells. The reporter construct contained the lacZ gene under the control of the nuclear factor of activated T cells (NF-AT) element of the human interleukin ... More
The role of the membrane domain in the regulated degradation of 3-hydroxy-3-methylglutaryl coenzyme A reductase.
AuthorsChun KT, Simoni RD
JournalJ Biol Chem
PubMed ID1740463
We have constructed a series of mutations in the membrane and linker domains of Syrian hamster 3-hydroxy-3-methylglutaryl-(HMG) CoA reductase in order to determine the regions critical for the regulated degradation of the enzyme. In transfected Chinese hamster ovary cells, we have expressed a fusion protein, HMGal, which consists of the ... More
Transfection activity of binary mixtures of cationic o-substituted phosphatidylcholine derivatives: the hydrophobic core strongly modulates physical properties and DNA delivery efficacy.
AuthorsWang L, Koynova R, Parikh H, MacDonald RC
JournalBiophys J
PubMed ID16935955
A combination of two cationic lipid derivatives having the same headgroup but tails of different chain lengths has been shown to have considerably different transfection activity than do the separate molecules. Such findings point to the importance of investigating the hydrophobic portions of cationic amphiphiles. Hence, we have synthesized a ... More
A novel fluorescence-based system for assaying and separating live cells according to VDJ recombinase activity.
AuthorsYancopoulos GD, Nolan GP, Pollock R, Prockop S, Li SC, Herzenberg LA, Alt FW
JournalMol Cell Biol
PubMed ID2320007
We describe two retroviral vector-based recombination substrate systems designed to assay for lymphoid VDJ recombinase activity in cultured cells. Both substrates incorporate a constitutive dominant marker gene (the simian virus promoter-driven neo gene) to allow selection of cells that stably integrate the substrate. Both substrates also include a second marker ... More
U937 cells deprived of endogenous annexin 1 demonstrate an increased PLA2 activity.
AuthorsSolito E, Raguenes-Nicol C, de Coupade C, Bisagni-Faure A, Russo-Marie F
JournalBr J Pharmacol
PubMed ID9756383
Annexin 1 (An 1), a phospholipid and calcium binding protein, is strongly expressed in differentiated U 937 cells. In attempting to correlate the expression of An 1 with phospholipase A2 (PLA2) activity, U 937 cells were stably transfected both with a Sense and Antisense cDNA for An 1. PLA2 activity ... More