TLR9 signals after translocating from the ER to CpG DNA in the lysosome.
AuthorsLatz E, Schoenemeyer A, Visintin A, Fitzgerald KA, Monks BG, Knetter CF, Lien E, Nilsen NJ, Espevik T, Golenbock DT
JournalNat Immunol
PubMed ID14716310
'Microbial DNA sequences containing unmethylated CpG dinucleotides activate Toll-like receptor 9 (TLR9). We have found that TLR9 is localized to the endoplasmic reticulum (ER) of dendritic cells (DCs) and macrophages. Because there is no precedent for immune receptor signaling in the ER, we investigated how TLR9 is activated. We show ... More
Targeted gene transfer to hepatocellular carcinoma cells in vitro using a novel monoclonal antibody-based gene delivery system.
AuthorsMohr L, Schauer JI, Boutin RH, Moradpour D, Wands JR
JournalHepatology
PubMed ID9862854
'Gene therapy approaches for the treatment of malignant tumors will require high-level expression of therapeutic genes in tumors compared with normal tissues. This may be achieved either by targeted gene delivery to tumor cells or by the use of tumor-specific promoters. Here, we describe the use of a novel conjugate ... More
Ethidium monoazide for DNA-based differentiation of viable and dead bacteria by 5'-nuclease PCR.
AuthorsNogva HK, Drømtorp SM, Nissen H, Rudi K
JournalBiotechniques
PubMed ID12703305
'PCR techniques have significantly improved the detection and identification of bacterial pathogens. Even so, the lack of differentiation between DNA from viable and dead cells is one of the major challenges for diagnostic DNA-based methods. Certain nucleic acid-binding dyes can selectively enter dead bacteria and subsequently be covalently linked to ... More
Effect of various short-term storage methods on viability of cancellous bone fragments.
AuthorsMcAnulty JF
JournalAm J Vet Res
PubMed ID9918149
'OBJECTIVE: To determine effects of various storage methods on ex vivo viability of cancellous bone fragments. SAMPLE POPULATION: Cancellous bone fragments obtained from 4 New Zealand White rabbits. PROCEDURE: Cancellous bone fragments were stored for 3 hours on ice in 1 of 5 preservation solutions or in 0.9% NaCl. Fragments ... More
Analysis of HCV-specific T cells by flow cytometry.
AuthorsShiina M, Rehermann B,
JournalMethods Mol Biol
PubMed ID19009279
'Flow cytometry has become an essential research tool because of the increase in the number of its {applications.} The development of an increasing number of monoclonal antibodies (mAbs) and fluorochromes, and of instruments capable of multicolor detection, allows the acquisition of a large amount of phenotypic and functional information in ... More
A hydrophobic protein, chargerin II, purified from rat liver mitochondria is encoded in the unidentified reading frame A6L of mitochondrial DNA.
AuthorsHiguti T, Negama T, Takigawa M, Uchida J, Yamane T, Asai T, Tani I, Oeda K, Shimizu M, Nakamura K
JournalJ Biol Chem
PubMed ID3360805
'Previous studies showed that a hydrophobic protein called chargerin II may have a key role in energy transduction of oxidative phosphorylation, since antibody against chargerin II labeled with monoazide ethidium inhibited ATP synthesis, ATP-Pi exchange, and reversed electron flow from succinate to NAD coupled with succinate oxidation by O2. In ... More
Use of a photolabeling technique to identify nonviable cells in fixed homologous or heterologous cell populations.
AuthorsRiedy MC, Muirhead KA, Jensen CP, Stewart CC
JournalCytometry
PubMed ID2049970
'Flow cytometric determination of viable versus nonviable cells in fixed samples can be accomplished by utilizing the irreversible binding of photoactivated ethidium monoazide (EMA). EMA is a positively charged molecule which is excluded by cells with intact membranes (viable cells), included by cells with damaged membranes, and can be photochemically ... More
Direct in vitro photoaffinity labeling of DNA with daunorubicin, adriamycin, and rubidazone.
AuthorsDaugherty JP, Hixon SC, Yielding KL
JournalBiochim Biophys Acta
PubMed ID508759
'Irradiation by daylight fluorescent lamps, of a physical complex of duplex DNA with either daunorubicin, adriamycin, or rubidazone produced covalent adducts to DNA. The photoincorporated drug could not be extracted by phenol extraction which removed 99% of the non-photolyzed drug from its physical complex with DNA. The photoadduct was also ... More
Signal- and energy-dependent nuclear transport of SV40 Vp3 by isolated nuclei. Establishment of a filtration assay for nuclear protein import.
AuthorsDean DA, Kasamatsu H
JournalJ Biol Chem
PubMed ID8106464
'Nuclear transport signal (NTS)-containing proteins are transported into the nucleus through the nuclear pore complex by a mechanism that is not well understood. To better characterize the mechanisms of transport, we have established an homologous in vitro system using an NTS-containing structural protein of simian virus 40 (SV40) and isolated ... More
Production of frameshift mutations in Salmonella by phenanthridinium derivatives: enzymatic activation and photoaffinity labeling.
AuthorsFukunaga M, Cox BA, von Sprecken RS, Yielding LW
JournalMutat Res
PubMed ID6374441
'The effect of metabolic activation on the mutagenic potential of some phenanthridinium compounds was examined in Salmonella typhimurium strains TA1538 and TA1978 . All of the compounds tested were mutagenic in TA1538, a DNA excision-repair-deficient strain, when metabolizing enzymes were included in the assay. Reversions were not detected when these ... More
Detection of DNA fragmentation in apoptosis: application of in situ nick translation to cell culture systems and tissue sections.
AuthorsGold R, Schmied M, Rothe G, Zischler H, Breitschopf H, Wekerle H, Lassmann H
JournalJ Histochem Cytochem
PubMed ID8515045
'Since DNA fragmentation is a key feature of programmed cell death (PCD) and also occurs in certain stages of necrosis, we have adapted the methodology of in situ nick-translation (ISNT) to detect DNA fragmentation on a single-cell level. We first established the technique for cell preparations. Apoptosis was induced by ... More
Analysis of in vivo expressed genes in Mannheimia haemolytica A1.
AuthorsLo RY, Sathiamoorthy S, Shewen PE
JournalFEMS Microbiol Lett
PubMed ID16984402
'The expression of Mannheimia haemolytica A1 genes during in vivo growth was examined by reverse transcriptase-polymerase chain reaction (RT-PCR) using total RNA extracted directly from M. haemolytica A1 recovered from pneumonic lungs of cattle. Primers specific for three groups of genes were used. Group 1 includes virulence-related genes: lktC, tbpB, ... More
Comparative binding of ethidium and three azido analogs to dinucleotides: affinity and intercalation geometry. A 1H NMR and visible spectroscopy study.
AuthorsLaugaa P, Delbarre A, Le Pecq JB, Roques BP
JournalEur J Biochem
PubMed ID6861758
'Geometrical and thermodynamic information has been obtained from theoretical analysis of both visible and 1H-NMR spectroscopic binding isotherms of ethidium and three photoactivable derivatives (8-azido-ethidium, 3-azido-ethidium and 3,8-diazido-ethidium) to self-complementary ribodinucleosides. The following results have been obtained. 1. Interaction with pyrimidine(3-5'')purine sequences is well accounted for by multicomponent equilibria involving ... More
Reversal of immunosuppression inducible through ultraviolet-exposed skin by in vivo anti-CD11b treatment.
AuthorsHammerberg C, Duraiswamy N, Cooper KD
JournalJ Immunol
PubMed ID8955170
'In both human in vitro models and murine in vivo adoptive transfer studies, UV-induced class II MHC+ CD11b+ leukocytes that infiltrate the epidermis appear to mediate UV-induced immunosuppression. In the present study, their role is further probed using an anti-CD11b mAb (clone 5C6), which is effective in vivo in blocking ... More
Histochemical applications of two phenanthridinium compounds.
AuthorsCox BA, Shackleford JM, Yielding LW
JournalStain Technol
PubMed ID6183795
'The fluorescent compounds ethidium monoazide and ethidium bromide were found to react intensely with nucleic acids of fixed, paraffin embedded tissues of rat and mouse. For routine staining, 10(-5) M solutions of ethidium bromide and its monoazide analogue were virtually identical in their reactions. Fresh frozen sections of the tissues ... More
Activation-induced deaminase cloning, localization, and protein extraction from young VH-mutant rabbit appendix.
'Studies in mouse, human, and chicken suggest that activation-induced deaminase (AID) is involved in three known processes leading to antibody diversification: somatic hypermutation, gene conversion, and class-switch recombination. Developing rabbit appendix provides a particularly good site for studying all three of these B cell maturation events. We report here successful ... More
Conformational coupling in H+-pumps and ATP synthesis--its analysis with anisotropic inhibitors of energy transduction in oxidative phosphorylation.
AuthorsHiguti T
JournalMol Cell Biochem
PubMed ID6323966
'The analysis of anisotropic inhibitor-induced phenomena in mitochondria revealed that two kinds of negative charges are generated near surface of the C-side of mitochondrial inner membranes in the energized state, on the redox complexes (I, III & IV) and F0, respectively, and that positively charged anisotropic inhibitors (AI+) inhibit energy ... More
Apoptosis in hematopoietic cells is associated with an extensive decrease in cellular phosphotyrosine content that can be inhibited by the tyrosine phosphatase antagonist pervanadate.
AuthorsLund-Johansen F, Frey T, Ledbetter JA, Thompson PA
JournalCytometry
PubMed ID8891448
'In the present study, we investigated whether apoptosis in hematopoietic cells is associated with changes in cellular phosphotyrosine content. Murine thymocytes and B cells, human leukemia cells, and normal peripheral blood leukocytes were induced to undergo apoptosis by treatment with specific stimuli or by incubation in growth factor-deprived medium. Multiparameter ... More
Variable immunodominance hierarchies for H2-M3-restricted N-formyl peptides following bacterial infection.
AuthorsKerksiek KM, Busch DH, Pamer EG
JournalJ Immunol
PubMed ID11145694
'H2-M3-restricted presentation of N-formyl methionine (f-Met) peptides to CD8(+) T cells provides a mechanism for selective recognition of bacterial infection. In this report we demonstrate that Listeria monocytogenes infection induces distinct CD8(+) T cell populations specific for each of the known Listeria-derived formyl methionine peptides presented by M3. The sum ... More
Comparison of petite induction in yeast by acridines, ethidium and their photoaffinity probes.
AuthorsFukunaga M, Yielding LW, Firth WJ, Yielding KL
JournalMutat Res
PubMed ID7022173
'The production of petite mutations by different acridine analogs was studied in Saccharomyces cerevisiae. Compounds with amino substituents at the 2 and 3 positions of the acridine nucleus and methylation at position 10 were effective for petite induction in growing cells but not in resting cells, while those with chloro, ... More
Covalent binding of ethidium azide analogs to Salmonella DNA in vivo: competition by ethidium bromide.
AuthorsYielding LW, Graves DE, Brown BR, Yielding KL
JournalBiochem Biophys Res Commun
PubMed ID444231
Structure-function characterization of phenanthridinium compounds as mutagens in Salmonella.
AuthorsFukunaga M, Yielding LW
JournalMutat Res
PubMed ID6348533
'The mutagenic activity of some phenanthridinium compounds was examined by using Salmonella typhimurium strain TA98. Microsomal enzyme activation of the compounds was necessary for the detection of frameshift mutagenesis. Amino and/or azido functions at both R3 and R8 were structural requisites for significant mutagenic activity, although mutagenicity was severely reduced ... More
Binding of ethidium monoazide to the chromatin in human lymphocytes.
AuthorsCantrell CE, Yielding KL
JournalBiochim Biophys Acta
PubMed ID6157417
'The azide analog of [14C]ethidium bromide was mixed with lymphocytes and photolyzed with visible light. The distribution of azide in the chromatin fraction was found to be 55% in DNA, 28% in protein and 16% in RNA. Label in the DNA portion was found to be almost exclusively in the ... More
7-Azido-actinomycin D: a photoaffinity probe of the sequence specificity of DNA binding by actinomycin D.
AuthorsRill RL, Marsch GA, Graves DE
JournalJ Biomol Struct Dyn
PubMed ID2627300
'Actinomycin D (ActD) is a DNA-binding antitumor antibiotic that appears to act in vivo by inhibiting RNA polymerase. The mechanism of DNA binding of ActD has attracted much attention because of its strong preference for 5''-dGpdC-3'' sequences. Binding is thought to involve intercalation of the tricyclic aromatic phenoxazone ring into ... More
Purified hydrophobic proteins, chargerins, are essential for energy transduction in oxidative phosphorylation.
AuthorsHiguti T, Takigawa M, Kotera Y, Oka H, Uchida J, Arakaki R, Fujita T, Ogawa T
JournalProc Natl Acad Sci U S A
PubMed ID2858097
'Studies on anisotropic inhibitors, a unique type of inhibitor of energy transduction in oxidative phosphorylation, suggested that redox reactions generate two kinds of negative charges on the outer surface of mitochondrial inner membranes, on redox complexes and on F0, and that the inhibitors inhibit energy transduction by binding to these ... More
Flow cytometric analysis of goat milk lymphocytes: subpopulations and adhesion molecule expression.
AuthorsGuiguen F, Greenland T, Pardo E, Panaye G, Mornex JF
JournalVet Immunol Immunopathol
PubMed ID8941978
'Cytometric analysis of cells in milk from healthy goats was achieved after elimination of interfering signals from debris and dead cells by irreversible staining with ethidium monoazide. Some 61% of milk lymphocytes are CD8+ T cells, 17% are CD4+ and about 20% of these express class II antigens: less than ... More
Kinetoplasts play an important role in the drug responses of Trypanosoma brucei.
AuthorsAgbe A, Yielding KL
JournalJ Parasitol
PubMed ID8544073
'Trypanosoma brucei E164 and a dyskinetoplastic derivative, Dysk164, were injected into mice that were treated subsequently with methylglyoxal-bis-guanylhydrazone, berenil, ethidium bromide, and acriflavine. Additionally, parasites were photoaffinity labeled with ethidium monoazide to effect covalent drug attachment prior to injection into animals. In all cases, killing of animals with E164 was ... More
Flow cytometric evaluation of the acrosome reaction of human spermatozoa: a new method using a photoactivated supravital stain.
AuthorsHenley N, Baron C, Roberts KD
JournalInt J Androl
PubMed ID7517916
'A flow cytometric assay using a double-stain method for the measurement of the acrosome reaction of human spermatozoa is described. The use of a stable photoactivated stain, ethidium monoazide, allowed evaluation of the viability of spermatozoa. This stain was more stable in fixed samples than propidium iodide, which is not ... More
Monoazido analog of ethidium as a chromatin probe: binding to DNA.
AuthorsCoffman GL, Yielding LW, Yielding L
JournalBiopolymers
PubMed ID6733248
Ethidium binding to deoxyribonucleic acid: spectrophotometric analysis of analogs with amino, azido, and hydrogen substituents.
AuthorsYielding LW, Yielding KL, Donoghue JE
JournalBiopolymers
PubMed ID6365193
Efficiency of photolytic binding of ethidium monoazide to nucleic acids and synthetic polynucleotides.
AuthorsCantrell CE, Yielding KL, Pruitt KM
JournalMol Pharmacol
PubMed ID572915
DNA excision repair in lymphocytes following photoaffinity labeling with ethidium monoazide.
AuthorsCantrell CE, Yielding KL
JournalPhotochem Photobiol
PubMed ID7220610
The interaction of 8-azidoethidium and tRNA.
AuthorsBertrand NJ, Kearns DR
JournalPhotochem Photobiol
PubMed ID7051058
Repair synthesis in human lymphocytes provoked by photolysis of ethidium azide.
AuthorsCantrell CE, Yielding KL
JournalPhotochem Photobiol
PubMed ID866451
Selective covalent binding of an ethidium analog to mitochondrial DNA with production of petite mutants in yeast by photoaffinity labelling.
AuthorsHixon SC, White WE, Yielding KL
JournalJ Mol Biol
PubMed ID1095756
Efficiency of photoaffinity labeling DNA homopolymers and copolymers with ethidium monoazide.
AuthorsDannelley JM, Boyce L, Gaubatz JW
JournalPhotochem Photobiol
PubMed ID3952163
Bypass by photoaffinity labeling of blocked metabolic activation of ethidium: confirmation of the role for covalent ethidium attachment in mitochondrial mutagenesis.
AuthorsHixon SC, White WE, Yielding KL
JournalBiochem Biophys Res Commun
PubMed ID1100050
Ethidium bromide and its photoreactive analogues: spectroscopic analysis of deoxyribonucleic acid binding properties.
AuthorsGraves DE, Watkins CL, Yielding LW
JournalBiochemistry
PubMed ID7225364
Nuclear mutations and mitotic recombination in Saccharomyces by light-activated ethidium azides.
AuthorsHixon SC, Burnham D
JournalMutat Res
PubMed ID379634
Cross-linking techniques.
AuthorsBäumert HG, Fasold H
JournalMethods Enzymol
PubMed ID2546017
Possible errors in the interpretation of ethidium bromide and PicoGreen DNA staining results from ethidium monoazide-treated DNA.
AuthorsHein I, Flekna G, Wagner M, Nocker A, Camper AK
JournalAppl Environ Microbiol
PubMed ID17021244
Intracellular binding of ethidium studied by photoaffinity labeling in vivo.
AuthorsFukunaga M, Yielding KL
JournalBiochim Biophys Acta
PubMed ID378267
The azide analog of 14C-labeled ethidium bromide was mixed with yeast cells and when photolyzed by visible light, formed covalent complexes with all yeast cell organelles. The 14C counts were found in DNA, RNA and protein of yeast subcellular fractions, illustrating the complexity of binding of a drug which appears ... More
Cross-linking of double-helical nucleic acids with a photoreactive analogue of ethidium.
AuthorsWoolley P, Dohrmann S
JournalBiochemistry
PubMed ID6882746
Intercalation of the ethidium analogue 3,5-diazido-5-ethyl-6-phenylphenanthridinium into double helices followed by irradiation with blue or ultraviolet light results in cross-linking between the two strands with an efficiency around 30% for DNA, RNA, and DNA-RNA hybrids. Details of this reaction and a convenient synthesis of the ethidium analogue are described. Stable ... More
Antagonism by propidium of petite induction by ethidium and ethidium azide in Saccharomyces cerevisiae.
AuthorsFukunaga M, Yielding KL
JournalMutat Res
PubMed ID386104
Propidium, a phenanthridinium dye similar to ethidium, did not induce petite mutations in non-growing yeast cells in contrast to ethidium. Combined exposure to ethidium and an excess of propidium for periods up to 2 h resulted in the expected petite induction expressed after subsequent plating on growth medium. As incubation ... More
Spectroscopic properties of ethidium monoazide: a fluorescent photoaffinity label for nucleic acids.
AuthorsBolton PH, Kearns DR
JournalNucleic Acids Res
PubMed ID745997
The non-covalent binding of ethidium monoazide to nucleic acids is entirely analogous to that of ethidium (binding constant approximately 2-3 X 10(5) M). The ethidium monoazide can be photochemically covalently linked to nucleic acids in high yield, up to 75%, by long wavelength light. The fluorescence of ethidium monoazide and ... More
Photoaffinity labeling of a mitochondrial hydrophobic protein by an anisotropic inhibitor of energy transduction in oxidative phosphorylation.
AuthorsHiguti T, Ohe T, Arakaki N, Kotera Y
JournalJ Biol Chem
PubMed ID7275983
The monoazide derivative of ethidium, the parent compound of which is an anisotropic inhibitor of energy transduction in oxidative phosphorylation, was synthesized and shown to be useful as a photoaffinity probe. Results showed that monoazide ethidium specifically binds to a hydrophobic protein of mitochondria (with an apparent molecular weight of ... More
[Structure-function characterization of phenanthridinium compounds as mutagens in salmonella and yeast]
AuthorsFukunaga M, Matsumoto N, Matsushita T, Mizuguchi Y
JournalJ UOEH
PubMed ID6385182
The relation between the mutagenic activities and chemical structure of phenanthridinium derivatives were tested by using Salmonella typhimurium strain TA 98 and yeast Saccharomyces cerevisiae. The 3,8-diamino analog and amino-azido isomers caused frameshift type mutation in Salmonella. However, mutagenicity was severely reduced for the diazido analog, and mutagenic activities of ... More
Deletion of mitochondrial genetic markers in yeast by ethidium and the photoaffinity probe, ethidium azide.
AuthorsFukunaga M, Yielding KL
JournalJpn J Med Sci Biol
PubMed ID393877
Induction of petite (cytoplasmic-respiration-deficient, rho-,rho-) mutations in yeast and deletion of mitochondrial drug-resistance genetic markers were compared after after treatment with ethidium and the corresponding photoaffinity probe, ethidium azide. Deletion of mitochondrial drug-resistance markers for chloramphenicol, erythromycin and oligomycin in these petite mutants was observed during prolonged treatment times with ... More
Antitrypanosomal action enhanced by photoaffinity labeling with ethidium analogs.
The trypanocidal activity of photoreactive azido analogs of ethidium was tested to determine the suitability of using such compounds as in vivo probes to study the mechanism of the antitrypanosomal activity of ethidium. Eight ethidium analogs, including three nonphotoreactive compounds, were tested for their ability to kill T. brucei both ... More
Synthesis, separation and characterization of the mono- and diazide analogs of ethidium bromide.
AuthorsGraves DE, Yielding LW, Watkins CL, Yielding KL
JournalBiochim Biophys Acta
PubMed ID911852
Ethidium bromide is used to characterize nucleic acid secondary and tertiary structural properties and the biological consequences of drug interactions. The mono- and diazido analogs of ethidium have proven valuable as photoaffinity probes in chemical and biological studies on nucleic acids, since they render the ethidium-nucleic acid interaction covalent. Although ... More
Use of ethidium monoazide and PCR in combination for quantification of viable and dead cells in complex samples.
AuthorsRudi K, Moen B, Drømtorp SM, Holck AL,
JournalAppl Environ Microbiol
PubMed ID15691961
The distinction between viable and dead cells is a major issue in many aspects of biological research. The current technologies for determining viable versus dead cells cannot readily be used for quantitative differentiation of specific cells in mixed populations. This is a serious limitation. We have solved this problem by ... More
Novel pentablock copolymers for selective gene delivery to cancer cells.
AuthorsZhang B, Kanapathipillai M, Bisso P, Mallapragada S,
JournalPharm Res
PubMed ID19142716
PURPOSE: In this study, the novel poly(diethylaminoethylmethacrylate) (PDEAEM)/Pluronic F127 pentablock copolymers were found to be able to mediate high-efficiency transfection of human epithelial ovarian carcinoma (SKOV3) cell line while showing significantly lower efficacy in human epithelial retinal (ARPE-19) cell line and Swiss Mouse Fibroblast (3T3) cell line. METHODS: The intracellular ... More
Comparison of apoptosis and mortality measurements in peripheral blood mononuclear cells (PBMCs) using multiple methods.
Death through apoptosis is the main process by which aged cells that have lost their function are eliminated. Apoptotic cells are usually detected microscopically by changes in their morphology. However, determination of early apoptotic events is important for in vitro (and ex vivo) studies. The main objective of the present ... More
Transfer of high copy number plasmid into mammalian cells by calcium phosphate transfection.
AuthorsBatard P, Jordan M, Wurm F
JournalGene
PubMed ID11404003
Using flow cytometry, single cell sorting, confocal microscopy and fluorescent plasmids, a thorough study of DNA uptake, DNA fate and DNA expression in mammalian cells transfected with the widely used calcium-phosphate precipitation method was executed. We show for the first time that up to 100,000 plasmid molecules can be delivered ... More
Gene inactivation by multiphoton-targeted photochemistry.
AuthorsBerns MW, Wang Z, Dunn A, Wallace V, Venugopalan V
JournalProc Natl Acad Sci U S A
PubMed ID10944219
Multiphoton-targeted photochemistry was used to selectively inactivate the expression of genes in vertebrate cells. A membrane permeable DNA-associating vital dye, ethidium bromide monoacetate (visible wavelength single photon absorption peak at 530 nm) was used to photosensitize chromosomes in dividing cells. A 100-ps infrared laser beam operating at 1.06 microns was ... More
Photofootprinting of drug-binding sites on DNA using diazo- and azido-9-aminoacridine derivatives.
AuthorsJeppesen C, Nielsen PE
JournalEur J Biochem
PubMed ID2472274
It is demonstrated that DNA photofootprinting analysis of the intercalating depsipeptide echinomycin, and the minor groove-binders distamicyn, 4',6-diamidino-2-phenylindole (DAPI) and Hoechst 33258 can be performed using 9-[6-(2-diazocyclopentadienylcarbonyloxy)hexylamino]acridine (DHA) [Nielsen et al. (1988) Nucleic Acids Res. 16, 3877-3888] or 2-methoxy-6-azido-9-aminoacridine (MAA) [Jeppesen et al. (1988) Nucleic Acids Res. 16, 5755-5770]. Both ... More
Detection of dead cells and measurement of cell killing by flow cytometry.
AuthorsKing MA
JournalJ Immunol Methods
PubMed ID10986413
A flow cytometer can quickly perform numerous quantitative, sensitive measurements on each individual cell within a large, heterogeneous population. The modern commercially available analytical instruments, which can be found in most hospitals, pathology laboratories, and cell biology research laboratories in the industrially developed countries, can now routinely measure fluorescence simultaneously ... More
Subcellular localization of photoreactive ethidium analogs in Trypanosoma brucei by fluorescence microscopy.
AuthorsCox BA, Yielding LW, Yielding KL
JournalJ Parasitol
PubMed ID6512635
To identify the in vivo targets of the trypanocide, ethidium bromide, the fluorescent staining of T. brucei was examined for a series of ethidium analogs using fluorescence microscopy. Determination of the biological targets for most drugs is limited by the reversible nature of their interactions. To overcome this limitation, photoaffinity ... More
Production of frameshift mutations in Salmonella by a light sensitive azide analog of ethidium.
AuthorsYielding LW, White WE, Yielding KL
JournalMutat Res
PubMed ID772423
Frameshift mutations have been produced in specific repair-negative Salmonella tester strains by photoaffinity labeling technique using ethidium azide. Reversions requiring a +1 addition or a -2 deletion were specially sensitive. Mutagenesis was reduced by the simultaneous addition of non-mutagenic ethidium bromide, and was prevented by photolysis of the azide prior ... More
Flow cytometric assay for the measurement of human bone marrow phenotype, function and cell cycle.
AuthorsLund-Johansen F, Bjerknes R, Laerum OD
JournalCytometry
PubMed ID2379452
A flow cytometric assay for the measurement of human bone marrow and blood leukocyte antigen expression, phagocytosis, and proliferation is described. Subpopulations of leukocytes were identified by their light scatter characteristics, and the expression of a myeloid differentiation antigen (designated CDw65) determined following incubation with CDw65 specific fluorescein-isothiocyanate (FITC) conjugated ... More
Comparative studies of the binding of ethidium bromide and its photoreactive analogues to nucleic acids by fluorescence and rapid kinetics.
AuthorsGarland F, Graves DE, Yielding LW, Cheung HC
JournalBiochemistry
PubMed ID7190829
The binding studies of the interaction of ethidium bromide with DNA have been hampered by its reversibility, which prevents direct isolation and thus characterization of the complex. The recent development of photoaffinity labeling has provided a means to circumvent this problem. However, to be useful as a probe for the ... More
Petite induction in Saccharomyces cerevisiae by ethidium analogs: distinction between resting and growing cells.
AuthorsFukunaga M, Yielding LW, Firth WJ, Yielding KL
JournalMutat Res
PubMed ID6993943
The importance of specific substituents, especially amino azide groups, for ethidium induction of petites was evaluated in resting and dividing cells of Saccharomyces cerevisiae through the study of a series of ethidium analogs. The structural requirements in resting and growing cells were found to be different, suggesting that at least ... More
Cytotoxicity and SOS-inducing ability of ethidium and photoactivable analogs on E. coli ethidium-bromide-sensitive (Ebs) strains.
AuthorsLambert B, Laugâa P, Roques BP, le Pecq JB
JournalMutat Res
PubMed ID2946948
In a recently-characterized ethidium-bromide-sensitive E. coli strain, DNA appears to be much more accessible to DNA-binding agents. This strain therefore appears to be of interest for studying the mutagenic properties of chemicals. For this purpose, a series of ethidium-sensitive E. coli strains (Ebs) with normal and defective DNA-repair capacity was ... More
Use of diazido ethidium bromide as a specific probe for mitochondrial functions.
AuthorsBastos RN
JournalJ Biol Chem
PubMed ID126240
The diazido derivative of ethidium bromide has been synthesized as a potential photoaffinity label and shown to be at least as effective as a mitochondrial mutagen as the parent compound, with a similar mode of action. Exposure of mitochondria of Saccharomyces cerevisiae to the compound, followed by ultraviolet-irradiation, which converts ... More
Clinical applications of flow cytometry. Immunologic methods for measuring cell membrane and cytoplasmic antigens.
AuthorsStewart CC
JournalCancer
PubMed ID1540893
Flow cytometric analysis is the method of choice for classifying leukemia and lymphomas. When combined with other clinical and morphometric data, a definitive diagnosis can be made in the most difficult cases. By using a combination of specific antibodies that are each labeled with a different fluorochrome, panels can be ... More
Ethidium bromide enhancement of frameshift mutagenesis caused by photoactivatable ethidium analogs.
AuthorsYielding LW, Brown BR, Graves DE, Yielding KL
JournalMutat Res
PubMed ID392307
Ethidium azide analogs (3-amino-8-azido-ethidium monoazide and ethidium diazide) have been developed as photosensitive probes in order to analyze directly the reversible in vivo interactions of ethidium bromide. Our preliminary observations [11], relating the mutagenic potential of the monoazide analog of ethidium, have been extended and refined, using the highly purified ... More
Demonstration of specific high affinity binding sites in plasmid DNA by photoaffinity labeling with an ethidium analog.
We have used photoaffinity labeling of pBR322 DNA with 8-azido-3-amino-5-ethyl-6-phenylphenanthridinium chloride to demonstrate high affinity ethidium-binding sites. Plasmid equilibrated with as little as 1 drug/DNA molecule was photoactivated, freed of uncomplexed drug by ethanol precipitation, and subjected to restriction analysis. There was highly specific, rather than random, blockage of HhaI ... More
Agonists cause nuclear translocation of phosphatidylinositol 3-kinase gamma. A Gbetagamma-dependent pathway that requires the p110gamma amino terminus.
AuthorsMetjian A, Roll RL, Ma AD, Abrams CS
JournalJ Biol Chem
PubMed ID10488142
In hematopoietic cells, the signals initiated by activation of the phosphoinositide 3-kinase (PI3K) family have been implicated in cell proliferation and survival, membrane and cytoskeletal reorganization, chemotaxis, and the neutrophil respiratory burst. Of the four isoforms of human PI3K that phosphorylate phosphatidylinositol 4, 5-bisphosphate, only p110gamma (or PI3Kgamma) is associated ... More
Alkali lability and rapid initiation of excision repair following photoaffinity damage by ethidium azide.
AuthorsKulkarni MS, Yielding KL
JournalChem Biol Interact
PubMed ID3000636
DNA damage and repair provoked by ethidium azide (EA) photoaffinity labeling in mouse leukemia cells was studied by measuring sedimentation properties of nucleoids in neutral sucrose gradients, and it was found that the strand opening step was faster than that which followed damage of cells by ultraviolet (UV) light. The ... More
Structure-function characterization for ethidium photoaffinity labels as mutagens in Salmonella.
AuthorsYielding LW, Firth WJ
JournalMutat Res
PubMed ID6993933
The development of photoaffinity probes to characterize the binding process and subsequent biological activity of a drug has recently been emphasized by the synthesis of two ethidium azide analogs. The initial finding showed that one of the azido analogs, the 8-azido-3-amino derivative, was at least 40-fold more mutagenic and toxic ... More
Binding of ethidium to the nucleosome core particle. 2. Internal and external binding modes.
AuthorsMcMurray CT, Small EW, van Holde KE
JournalBiochemistry
PubMed ID2043608
We have previously reported that the binding of ethidium bromide to the nucleosome core particle results in a stepwise dissociation of the structure which involves the initial release of one copy each of H2A and H2B (McMurray & van Holde, 1986). In this report, we have examined the absorbance and ... More
Use of drug-specific antibodies to identify ethidium adducts produced in Trypanosoma brucei by photoaffinity labeling.
A photoreactive azido analog of the trypanocide ethidium bromide, 3-amino-8-azido-5-ethyl-6-phenylphenanthridinium chloride, attached covalently to calf thymus DNA (CT DNA) by photoaffinity labeling, was used to generate antibodies for the drug analog. The specificity of the antiserum was tested using enzyme-linked immunoadsorbant assays (ELISA) against immobilized antigen (photoaffinity labeled DNA) and ... More
A new cationic liposome for efficient gene delivery with serum into cultured human cells: a quantitative analysis using two independent fluorescent probes.
AuthorsSerikawa T, Suzuki N, Kikuchi H, Tanaka K, Kitagawa T
JournalBiochim Biophys Acta
PubMed ID11030599
Cationic liposomes are useful to transfer genes into eukaryotic cells in vitro and in vivo. However, liposomes with good transfection efficiency are often cytotoxic, and also require serum-free conditions for optimal activity. In this report, we describe a new formulation of cationic liposome containing DC-6-14, O,O'-ditetradecanoyl-N-(alpha-trimethylammonioacetyl)diethan olamine chloride, dioleoylphosphatidylethanolamine and ... More
Ethidium binding sites on plasmid DNA determined by photoaffinity labeling.
AuthorsHardwick JM, von Sprecken RS, Yielding KL, Yielding LW
JournalJ Biol Chem
PubMed ID6088528
Photoaffinity labeling of pBR322 with ethidium monoazide (8-azido-3-amino-5-ethyl-6-phenylphenanthridinium chloride) was used to provide evidence for the sequence specificity of ethidium binding to native DNA. DNA-drug interactions were examined at concentrations of eight covalently bound ethidium drugs per molecule of pBR322 (4363 base pairs). Restriction enzyme cutting was blocked by the ... More
Synergistic neurotoxicity of opioids and human immunodeficiency virus-1 Tat protein in striatal neurons in vitro.
AuthorsGurwell JA, Nath A, Sun Q, Zhang J, Martin KM, Chen Y, Hauser KF
JournalNeuroscience
PubMed ID11226693
Human immunodeficiency virus (HIV) infection selectively targets the striatum, a region rich in opioid receptor-expressing neural cells, resulting in gliosis and neuronal losses. Opioids can be neuroprotective or can promote neurodegeneration. To determine whether opioids modify the response of neurons to human immunodeficiency virus type 1 (HIV-1) Tat protein-induced neurotoxicity, ... More
Inhibition of the B to Z transition in poly(dGdC).poly(dGdC) by covalent attachment of ethidium: equilibrium studies.
AuthorsGilbert PL, Graves DE, Chaires JB
JournalBiochemistry
PubMed ID1932017
The effects of covalent modification of poly(dGdC).poly(dGdC) and poly(dGm5dC).poly(dGm5dC) by ethidium monoazide (a photoreactive analogue of ethidium) on the salt-induced B to Z transition are examined. Earlier studies have shown ethidium monoazide to bind DNA (in the absence of light) in a manner identical to that of the parent ethidium ... More
Inhibition of the B to Z transition in poly(dGdC).poly(dGdC) by covalent attachment of ethidium: kinetic studies.
AuthorsGilbert PL, Graves DE, Britt M, Chaires JB
JournalBiochemistry
PubMed ID1932018
The photoaffinity analogue ethidium monoazide was used to prepare samples of poly(dGdC).poly(dGdC) containing covalently attached ethidium. The effects of both noncovalently and covalently bound ethidium on the kinetics of the NaCl-induced B to Z transition in poly(dGdC).poly(dGdC) was examined using absorbance and fluorescence spectroscopy to monitor the reaction. Covalently and ... More
Covalent attachment of ethidium to DNA results in enhanced topoisomerase II-mediated DNA cleavage.
AuthorsMarx G, Zhou H, Graves DE, Osheroff N
JournalBiochemistry
PubMed ID9398321
The classic DNA intercalator, ethidium, was used to probe the effects of (i) intercalation and (ii) covalent modification of the DNA on the catalytic activity of topoisomerase II. Ethidium bromide, which binds reversibly to DNA via intercalation, does not stimulate topoisomerase II-mediated DNA cleavage at concentrations up to 100 microM, ... More
In vitro micronucleus scoring by flow cytometry: differential staining of micronuclei versus apoptotic and necrotic chromatin enhances assay reliability.
AuthorsAvlasevich SL, Bryce SM, Cairns SE, Dertinger SD
JournalEnviron Mol Mutagen
PubMed ID16180205
The in vitro micronucleus test has received considerable attention in recent years for its use in drug safety assessment and toxicological research. The less tedious nature of the assay relative to chromosome aberration analyses is a driving force, and explains why many chemical and drug safety programs have adopted the ... More
Comparison of cell viability probes compatible with fixation and permeabilization for combined surface and intracellular staining in flow cytometry.
AuthorsO'Brien MC, Bolton WE
JournalCytometry
PubMed ID7537649
Dead cells represent a significant source of interference in the flow cytometric analysis of viable cells primarily due to nonspecific uptake of probes, increased autofluorescence, and altered antigen expression and DNA content. Traditional methods of dead cell exclusion, based on light scatter or uptake of dyes such as propidium iodide ... More
In vitro micronucleus assay scored by flow cytometry provides a comprehensive evaluation of cytogenetic damage and cytotoxicity.
This laboratory has previously reported on the development of a flow cytometry-based method for scoring in vitro micronuclei in mouse lymphoma (L5178Y) cells [S.L. Avlasevich, S.M. Bryce, S.E. Cairns, S.D. Dertinger, In vitro micronucleus scoring by flow cytometry: differential staining of micronuclei versus apoptotic and necrotic chromatin enhances assay reliability, ... More
Photoaffinity approaches to determining the sequence selectivities of DNA-small molecule interactions: actinomycin D and ethidium.
AuthorsMarsch GA, Graves DE, Rill RL
JournalNucleic Acids Res
PubMed ID7739904
The DNA photoaffinity ligands, 7-azidoactinomycin D and 8-azidoethidium, form DNA adducts that cause chain cleavage upon treatment with piperidine. Chemical DNA sequencing techniques were used to detect covalent binding. The relative preferences for modifications of all possible sites defined by a base pair step (e.g. GC) were determined within all ... More
Mitosis enhances transgene expression of plasmid delivered by cationic liposomes.
AuthorsTseng WC, Haselton FR, Giorgio TD
JournalBiochim Biophys Acta
PubMed ID10209258
A critical requirement of gene therapy is expression of the delivered transgene. Transgene expression is facilitated by access to the transcription mechanism found primarily in the nucleus. Factors modulating the interactions between intracellular plasmid and nuclear access are not well understood. In this study, the effect of mitosis on transgene ... More
Site-selective photoaffinity labeling of the Torpedo californica nicotinic acetylcholine receptor by azide derivatives of ethidium bromide.
AuthorsPedersen SE
JournalMol Pharmacol
PubMed ID7838117
Three azido derivatives of ethidium bromide, a potent noncompetitive antagonist of the nicotinic acetylcholine receptor from Torpedo californica, were synthesized, namely 8-azido-ethidium chloride, 3-azido-ethidium chloride, and 3,8-diazido-ethidium chloride. These derivatives were tested for their ability to interact with the noncompetitive antagonist binding site and the acetylcholine binding sites on the ... More
Cationic albumin-conjugated pegylated nanoparticles allow gene delivery into brain tumors via intravenous administration.
AuthorsLu W, Sun Q, Wan J, She Z, Jiang XG
JournalCancer Res
PubMed ID17178885
Patients with malignant gliomas have a poor prognosis because these tumors do not respond well to conventional treatments. Studies of glioma xenografts suggest that they may be amenable to gene therapy with cytotoxic genes, such as the proapoptotic Apo2 ligand/tumor necrosis factor-related apoptosis-inducing ligand (Apo2L/TRAIL). Gene therapy of gliomas ideally ... More
A fluorescence photobleaching study of the microsecond reorientational motions of DNA.
AuthorsScalettar BA, Selvin PR, Axelrod D, Hearst JE, Klein MP
JournalBiophys J
PubMed ID2964258
We have conducted a polarized fluorescence photobleaching recovery (FPR) study of the rotational dynamics of ethidium azide labeled DNA. Polarized photobleaching experiments provide data on microsecond and millisecond molecular reorientation that complement the information available from nanosecond fluorescence depolarization studies. In polarized FPR experiments an anisotropic angular concentration of fluorophore ... More
A monoclonal antibody cytolytic to androgen independent DU145 and PC3 human prostatic carcinoma cells.
BACKGROUND: While a range of therapeutic products is available for androgen-dependent prostatic cancer, no specific intervention modality exists for androgen-independent prostatic cancer. The objective of this research was to explore whether epitopes exist on androgen-independent prostatic DU145 cancer cells, which could be susceptible to cytotoxic action of specific antibodies. METHODS: ... More
Transfection by cationic liposomes using simultaneous single cell measurements of plasmid delivery and transgene expression.
AuthorsTseng WC, Haselton FR, Giorgio TD
JournalJ Biol Chem
PubMed ID9325286
Cationic liposomes are potentially important gene transfer vehicles, although their application has been limited by relatively low efficiency of transgene expression. Single cell quantitative methods, such as those used in this study, should permit a more detailed understanding of the relationships between delivered plasmid and transgene expression. Intracellular plasmid delivery ... More
Human Fetal Progenitor Tenocytes for Regenerative Medicine.
Authors
JournalCell Transplant
PubMed ID26110286
Argovit™ silver nanoparticles to fight Huanglongbing disease in Mexican limes (Citrus aurantifolia Swingle).
Authors
JournalRSC Adv
PubMed ID35495993
Recruitment of highly cytotoxic CD8+ T cell receptors in mild SARS-CoV-2 infection.
Authors
JournalCell Rep
PubMed ID34968416
Platelet P-selectin initiates cross-presentation and dendritic cell differentiation in blood monocytes.
Authors
JournalSci Adv
PubMed ID32195350
High content drug screening for Fanconi anemia therapeutics.
Authors
JournalOrphanet J Rare Dis
PubMed ID32605631
Fate mapping of single NK cells identifies a type 1 innate lymphoid-like lineage that bridges innate and adaptive recognition of viral infection.
Authors
JournalImmunity
PubMed ID34437840
Evaluation of genotoxic potential of peptides used in nuclear medicine (PSMA -617 and -11, and ubiquicidine 29-41) using a flow-cytometric, semi-automated analysis of micronuclei frequency in cell cultures.
Authors
JournalToxicol Rep
PubMed ID32071884
Efficient differentiation of cardiomyocytes from human pluripotent stem cells with growth factors.
Authors
JournalMethods Mol Biol
PubMed ID25836579
A long non-coding RNA GATA6-AS1 adjacent to GATA6 is required for cardiomyocyte differentiation from human pluripotent stem cells.
Authors
JournalFASEB J
PubMed ID32888237
Programming thermoresponsiveness of NanoVelcro substrates enables effective purification of circulating tumor cells in lung cancer patients.