Protonation dynamics of the extracellular and cytoplasmic surface of bacteriorhodopsin in the purple membrane.
AuthorsNachliel E,Gutman M,Kiryati S,Dencher NA
JournalProceedings of the National Academy of Sciences of the United States of America
PubMed ID8855251
The dynamics of proton binding to the extracellular and the cytoplasmic surfaces of the purple membrane were measured by laser-induced proton pulses. Purple membranes, selectively labeled by fluorescein at Lys-129 of bacteriorhodopsin, were pulsed by protons released in the aqueous bulk from excited pyranine (8-hydroxy-1,3,6-pyrenetrisulfonate) and the reaction of protons ... More
The engineering of membrane-permeable peptides.
AuthorsCarrigan CN,Imperiali B
JournalAnalytical biochemistry
PubMed ID15907875
Sensitive fluorescence-based thermodynamic and kinetic measurements of DNA hybridization in solution.
AuthorsMorrison LE,Stols LM
JournalBiochemistry
PubMed ID8457571
Kinetic and thermodynamic constants associated with DNA hybridization were determined in solution using fluorescence measurements and complementary fluorophore-labeled oligomers. One oligomer was labeled with a 5'-terminal fluorescein, and the other was labeled with a 3'-terminal rhodamine. The juxtaposition of the two labels in double-stranded complexes results in a strong quenching ... More
Fluorescence resonance energy transfer analysis of the structure of the four-way DNA junction.
We have carried out fluorescence resonance energy transfer (FRET) measurements on four-way DNA junctions in order to analyze the global structure and its dependence on the concentration of several types of ions. A knowledge of the structure and its sensitivity to the solution environment is important for a full understanding ... More
Macrophage phagocytosis: use of fluorescence microscopy to distinguish between extracellular and intracellular bacteria.
AuthorsDrevets DA, Campbell PA
JournalJ Immunol Methods
PubMed ID1919019
One of the challenges of phagocytosis research is to differentiate bacteria adherent to a host cell from bacteria which the cell has internalized. To address this question, various techniques such as fluorescence microscopy, electron microscopy, and flow cytometry have been used. We have adapted a flow cytometric method (Fattorossi et ... More
Multiple binding sites of fluorescein isothiocyanate moieties on myoglobin: photophysical heterogeneity as revealed by ground- and excited-state spectroscopy.
AuthorsGrunwaldt G, Haebel S, Spitz C, Steup M, Menzel R
JournalJ Photochem Photobiol B
PubMed ID12167317
Fluorescein isothiocyanate (FITC)-myoglobin conjugates were synthesized with a binding stoichiometry of one to three fluorophores per protein. FITC binding sites were determined by matrix-assisted laser desorption-ionization time-of-flight mass spectrometry (MALDI-TOF MS). Five lysine residues and the N-terminal amino group were identified as preferential binding sites. The ground and excited-state absorption ... More
Lateral mobility of phospholipids in the external and internal leaflets of normal and hereditary spherocytic human erythrocytes.
AuthorsRimon G, Meyerstein N, Henis YI
JournalBiochim Biophys Acta
PubMed ID6466671
The lateral diffusion coefficients (D) and the mobile fractions of the fluorescent phospholipid N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)phosphatidylethanolamine (NBD-PE) and of membrane proteins labelled with fluorescein isothiocyanate, were measured by fluorescence photobleaching recovery on erythrocytes from healthy persons and from a hereditary spherocytosis patient. Measurements of lipid probe mobility were performed on ghosts labelled ... More
Regulation of vascular smooth muscle tone by N-terminal region of caldesmon. Possible role of tethering actin to myosin.
AuthorsLee YH, Gallant C, Guo H, Li Y, Wang CA, Morgan KG
JournalJ Biol Chem
PubMed ID10652307
'To assess the functional significance of tethering actin to myosin by caldesmon in the regulation of smooth muscle contraction, we investigated the effects of synthetic peptides, containing the myosin-binding sequences in the N-terminal region of caldesmon, on force directly recorded from single permeabilized smooth muscle cells of ferret portal vein. ... More
Monoclonal antibodies to NTF2 inhibit nuclear protein import by preventing nuclear translocation of the GTPase Ran.
AuthorsSteggerda SM, Black BE, Paschal BM
JournalMol Biol Cell
PubMed ID10679025
'Nuclear transport factor 2 (NTF2) is a soluble transport protein originally identified by its ability to stimulate nuclear localization signal (NLS)-dependent protein import in digitonin-permeabilized cells. NTF2 has been shown to bind nuclear pore complex proteins and the GDP form of Ran in vitro. Recently, it has been reported that ... More
Immunochemical and spectroscopic characterization of two fluorescein 5'-isothiocyanate labeling sites on Na+,K(+)-ATPase.
AuthorsAbbott AJ, Amler E, Ball WJ
JournalBiochemistry
PubMed ID1847073
'Fluorescein 5'-isothiocyanate (FITC) covalently modifies the Lys-501 residue of the catalytic (alpha) subunit of Na+,K(+)-ATPase and resides at a conformation-sensitive site in or near the ATP binding site. In these studies, FITC-directed antibodies which quench this hapten's fluorescence were used to infer the solvent accessibility of the enzyme-bound probe. These ... More
Fluorescein and tetramethyl rhodamine as haptens in enzyme immunohistochemistry.
AuthorsHaaijman JJ, Coolen J, Kröse CJ, Pronk GJ, Ming ZF
JournalHistochemistry
PubMed ID3522496
'Fluorescein (Fl) and tetramethyl rhodamine (Rh) were evaluated as possible candidates for a double hapten sandwich system in enzyme immunohistology. Monoclonal antibodies were raised against Fl and Rh. Their fine-specificity was tested with a competition-like assay. A pair of Mab''s was selected for immunohistology in which they functioned as a ... More
Resonance energy transfer between guanine nucleotide binding protein subunits and membrane lipids.
AuthorsRemmers AE, Neubig RR
JournalBiochemistry
PubMed ID7680231
'Resonance energy transfer was used to estimate the distances of closest approach between fluorescent labels on G protein subunits (alpha, beta, and gamma) and the phospholipid bilayer surface. Fluorescein-labeled alpha, beta, and gamma subunits were the energy transfer donors and hexadecylaminoeosin (HAE) in phospholipid vesicles was the acceptor. Bovine brain ... More
Time resolution of fluorescence changes observed in titrations of fluorescein 5'-isothiocyanate-modified Na,K-ATPase with monovalent cations.
AuthorsLin SH, Faller LD
JournalBiochemistry
PubMed ID8268167
'Equilibrium fluorometric titrations of fluorescein 5''-isothiocyanate-modified Na,K-ATPase with cations have usually been interpreted by assuming that an enhancement reports the conformational change from E2 to E1. We report time resolution of the fluorescence change into three phases when fluorophore-modified enzyme is mixed with the chloride salt of either sodium or ... More
Aggretin, a heterodimeric C-type lectin from Calloselasma rhodostoma (malayan pit viper), stimulates platelets by binding to alpha 2beta 1 integrin and glycoprotein Ib, activating Syk and phospholipase Cgamma 2, but does not involve the glycoprotein VI/Fc receptor gamma chain collagen receptor.
AuthorsNavdaev A, Clemetson JM, Polgar J, Kehrel BE, Glauner M, Magnenat E, Wells TN, Clemetson KJ
JournalJ Biol Chem
PubMed ID11287424
'Aggretin, a potent platelet activator, was isolated from Calloselasma rhodostoma venom, and 30-amino acid N-terminal sequences of both subunits were determined. Aggretin belongs to the heterodimeric snake C-type lectin family and is thought to activate platelets by binding to platelet glycoprotein alpha(2)beta(1). We now show that binding to glycoprotein (GP) ... More
Simultaneous flow cytometric detection of cellular c-myc protein, incorporated bromodeoxyuridine, and DNA.
AuthorsRosette CD, DeTeresa PS, Pallavicini MG
JournalCytometry
PubMed ID2111761
'We describe a multivariate flow cytometric technique for simultaneous analysis of specific nuclear protein, bromodeoxyuridine (BrdUrd) incorporated into DNA and DNA content in single cells in suspension. The procedure involves fixation of BrdUrd-exposed cells with paraformaldehyde, heat denaturation of cellular DNA, followed by sequential immunochemical reactions to label incorporated BrdUrd ... More
CD14-dependent internalization of bacterial lipopolysaccharide (LPS) is strongly influenced by LPS aggregation but not by cellular responses to LPS.
AuthorsKitchens RL, Munford RS
JournalJ Immunol
PubMed ID9469454
'We analyzed the impact of ligand aggregation and LPS-induced signaling on CD14-dependent LPS internalization kinetics in human monocytic THP-1 cells and murine macrophages. Using two independent methods, we found that the initial rate and extent of LPS internalization increased with LPS aggregate size. In the presence of LPS binding protein ... More
Distribution of microtubules and other cytoskeletal filaments during myotube elongation as revealed by fluorescence microscopy.
AuthorsSaitoh O, Arai T, Obinata T
JournalCell Tissue Res
PubMed ID3383210
'Distribution of microtubules and other cytoskeletal filaments in growing skeletal muscle cells (myotubes) was studied in vitro by fluorescence microscopy using fluorescin-labeled antibodies and phalloidin, a specific antiactin drug. In the distal elongating tips of myotubes, microtubules were the major cytoskeletal elements; actin and intermediate filaments were much less abundant. ... More
Regulation of alphaIIb beta3 function in human B lymphocytes.
AuthorsQi W, Loh E, Vilaire G, Bennett JS
JournalJ Biol Chem
PubMed ID9614143
'We studied the function of the platelet integrin alphaIIb beta3 using a B lymphocyte model in which alphaIIb beta3 can be induced to interact with fibrinogen using phorbol myristate acetate (PMA). To determine whether a G protein-coupled receptor could also activate alphaIIb beta3 in lymphocytes, we coexpressed the human formyl ... More
Functional studies of the domains of talin.
AuthorsNuckolls GH, Turner CE, Burridge K
JournalJ Cell Biol
PubMed ID2110569
'The protein talin has two domains of approximately 200 and 47 kD, which can be cleaved apart by a variety of proteases. To examine the function of these two structural domains of talin, we have digested purified talin with a calcium-dependent protease and separated the resulting fragments chromatographically. Both fragments ... More
Acid-induced conformational changes in phosphoglucose isomerase result in its increased cell surface association and deposition on fibronectin fibrils.
AuthorsAmraei M, Jia Z, Reboul P, Nabi IR
JournalJ Biol Chem
PubMed ID12888574
'Phosphoglucose isomerase (PGI) is a glycolytic enzyme that exhibits extracellular cytokine activity as autocrine motility factor, neuroleukin, and maturation factor and that has been recently implicated as an autoantigen in rheumatoid arthritis. In contrast to its receptor-mediated endocytosis at neutral pH, addition of 25 microg/ml of either Alexa 568- or ... More
TCR engagement of CD4+CD8+ thymocytes in vitro induces early aspects of positive selection, but not apoptosis.
AuthorsGroves T, Parsons M, Miyamoto NG, Guidos CJ
JournalJ Immunol
PubMed ID8977176
'Immature CD4/CD8 double-positive (DP) thymocytes expressing self MHC-restricted TCR are positively selected in response to TCR signals to survive and differentiate into functionally competent CD4 or CD8 single positive (SP) T cells. In contrast, DP precursors expressing autoreactive TCR are clonally deleted in response to TCR signals. We show here ... More
Fluorescence anisotropy studies of enzyme-substrate complex formation in stearoyl-ACP desaturase.
AuthorsHaas JA, Fox BG
JournalBiochemistry
PubMed ID12463745
'Stearoyl-acyl carrier protein Delta(9)-desaturase (delta9D) catalyzes regio- and stereospecific insertion of cis double bonds into acyl chains attached to acyl carrier protein. Steady-state and stopped-flow fluorescence anisotropy measurements using acylated forms of dansyl- and fluoresceinyl-ACPs revealed equilibrium dissociation constants and dissociation rate constants for 16:0-, 17:0-, and 18:0-ACPs with resting ... More
Absence of Na+,K(+)-ATPase regulation of endosomal acidification in K562 erythroleukemia cells. Analysis via inhibition of transferrin recycling by low temperatures.
AuthorsSipe DM, Jesurum A, Murphy RF
JournalJ Biol Chem
PubMed ID1847374
'Transferrin (Tf) acidification has been shown to be limited to pH 6 in murine Balb/c 3T3 fibroblasts, human A549 epidermoid carcinoma cells, and Chinese hamster ovary cells and is followed by alkalinization during recycling. In contrast, Tf acidification in the human erythroleukemic cell line K562 proceeds to below pH 5.5, ... More
Conformational changes of the nucleotide site of the plasma membrane Ca2+-ATPase probed by fluorescence quenching.
AuthorsFonseca MM, Scofano HM, Carvalho-Alves PC, Barrabin H, Mignaco JA
JournalBiochemistry
PubMed ID12044182
'Fluorescence quenching by the water-soluble ions I(-) and Cs(+) was used to probe solvent accessibility and polarity of the nucleotide/fluorescein isothiocyanate binding pocket of the purified soluble Ca(2+)-ATPase from plasma membranes. The E(1).Ca.CaM conformer was the least accessible state studied, presenting the lowest suppression constant (K(q)) for both I(-) (K(q) ... More
The nucleotide-binding site of the sarcoplasmic reticulum Ca-ATPase is conformationally altered in aged skeletal muscle.
AuthorsChen B, Jones TE, Bigelow DJ
JournalBiochemistry
PubMed ID10555971
'Cellular conditions in senescent skeletal muscle have been shown to result in the loss of conformational stability of the sarcoplasmic reticulum (SR) Ca-ATPase. To identify underlying structural features of age-modified Ca-ATPase, we have utilized the fluorescence properties of protein-bound probes to assess both local and global structure. We find conformational ... More
Kinetics of antigenic peptide binding to the class II major histocompatibility molecule I-Ad.
AuthorsTampé R, McConnell HM
JournalProc Natl Acad Sci U S A
PubMed ID2052549
'Using high-performance size-exclusion chromatography and fluorescence spectroscopy, we investigated the kinetics of fluorescent peptide reactions with detergent-solubilized I-Ad, a class II molecule of the mouse major histocompatibility complex. At pH 7.0 and 37 degrees C the half-time for the binding of a fluorescein-labeled synthetic peptide representing ovalbumin amino acids 323-339 ... More
Interaction of melittin with the (Na+ + K+)ATPase: evidence for a melittin-induced conformational change.
AuthorsCuppoletti J, Abbott AJ
JournalArch Biochem Biophys
PubMed ID2177321
'The (Na+ + K+)ATPase is inhibited by the bee venom polypeptide, melittin. KCl and NaCl protect the enzyme from melittin inhibition. Analysis of the K+ and Na+ protection against melittin inhibition suggested a kinetic model which was consistent with slowly reversible melittin binding, and mutually exclusive binding of melittin with ... More
Co-expression of chemotactic ligand receptors on human peripheral blood monocytes.
AuthorsOhura K, Katona IM, Wahl LM, Chenoweth DE, Wahl SM
JournalJ Immunol
PubMed ID2435802
'Directed migration of monocytes is dependent upon interaction of cell surface receptors and specific chemotactic ligands. To determine whether circulating human monocytes express multiple chemotactic ligand receptors or whether subpopulations of monocytes exist with a single receptor specificity, nonoverlapping fluorescent probes for two chemotactic ligands, N-formyl methionyl leucyl phenylalanine (FMLP) ... More
Location of the epidermal growth factor binding site on the EGF receptor. A resonance energy transfer study.
AuthorsCarraway KL, Koland JG, Cerione RA
JournalBiochemistry
PubMed ID2271554
'As a first step toward developing a structural map of key sites on the epidermal growth factor (EGF) receptor, we have used resonance energy transfer to measure the distance of closest approach between the receptor-bound growth factor molecule and lipid molecules at the surface of the plasma membrane. EGF, specifically ... More
Intersubunit communications in Escherichia coli cyclic AMP receptor protein: studies of the ligand binding domain.
AuthorsHeyduk E, Heyduk T, Lee JC
JournalBiochemistry
PubMed ID1314647
'Escherichia coli cAMP receptor protein (CRP) is a homodimer in which each subunit is composed of two domains. The C-terminal domain is responsible for DNA recognition, whereas the larger N-terminal domain is involved in cAMP binding. Biochemical and genetic evidence suggests that both intersubunit and interdomain interactions play important roles ... More
Modification of fibroblast surface amines alters receptor-mediated cell spreading on protein-coated substrata but not adsorptive endocytosis.
AuthorsAplin JD, Hughes RC
JournalJ Cell Sci
PubMed ID6118378
'Fluorescein isothiocyanate (FITC) and other anionic reagents specific for amine groups have previously been shown to inhibit the adhesion and spreading of cultured fibroblasts to fibronectin-coated surfaces (Butters, Devalia, Aplin & Hughes, 1980). Here it is demonstrated that a population of FITC-labelled cells can be separated using flow cytometry into ... More
Identification of arginyl residues located at the ATP binding site of sarcoplasmic reticulum Ca2+-ATPase. Modification with 1,2-cyclohexanedione.
AuthorsKimura K, Suzuki H, Daiho T, Yamasaki K, Kanazawa T
JournalJ Biol Chem
PubMed ID8910542
'Sarcoplasmic reticulum vesicles were treated with 1, 2-cyclohexanedione (CHD) in sodium borate (pH 8.0). The Ca2+-ATPase activity was completely inhibited. Inhibition of Mg.ATP and Mg.ADP binding to the high affinity ATP binding site as well as inhibition of phosphorylation with ATP occurred simultaneously with the inhibition of the Ca2+-ATPase activity. ... More
Covalent modification of G-actin by pyridoxal 5'-phosphate: polymerization properties and interaction with DNase I and myosin subfragment 1.
AuthorsCombeau C, Carlier MF
JournalBiochemistry
PubMed ID1731881
'Pyridoxal 5''-phosphate (PLP), a lysine-specific reagent, has been used to modify G-actin. At pH 7.5, PLP reacted with 1.7-2 lysines on G-actin. Limited proteolytic digestion experiments indicated that, in agreement with previous works, essentially lysine-61 was modified in a 1:1 fashion by PLP, other lysines being much less reactive. A ... More
Subunit interactions of GTP-binding proteins.
AuthorsHeithier H, Fröhlich M, Dees C, Baumann M, Häring M, Gierschik P, Schiltz E, Vaz WL, Hekman M, Helmreich EJ
JournalEur J Biochem
PubMed ID1312936
'Fluorescence energy transfer [cf. Förster, T. (1948) Ann. Phys. 6, 55-75] was tested for its suitability to study quantitative interactions of subunits of G0 with each other and these subunits or trimeric G0 with the beta 1-adrenoceptor in detergent micelles or after reconstitution into lipid vesicles [according to Feder, D., ... More
B lymphocytes express and lose syndecan at specific stages of differentiation.
AuthorsSanderson RD, Lalor P, Bernfield M
JournalCell Regul
PubMed ID2519615
'Lymphopoietic cells require interactions with bone marrow stroma for normal maturation and show changes in adhesion to matrix during their differentiation. Syndecan, a heparan sulfate-rich integral membrane proteoglycan, functions as a matrix receptor by binding cells to interstitial collagens, fibronectin, and thrombospondin. Therefore, we asked whether syndecan was present on ... More
Specific associations of fluorescent beta-2-microglobulin with cell surfaces. The affinity of different H-2 and HLA antigens for beta-2-microglobulin.
AuthorsHochman JH, Shimizu Y, DeMars R, Edidin M
JournalJ Immunol
PubMed ID2450918
'We prepared single-labeled FITC derivatives of beta-2-microglobulin (b2m) and examined their interactions with class I MHC Ag H chains on living cells. Human b2m was reacted with FITC under mild conditions and separated by hydroxylapatite chromatography into three peaks containing single labeled derivatives of b2m peaks A, B, and C, ... More
Detection of conformational changes in chloroplast coupling factor 1 by 8-anilino-1-naphthalene-sulphonate fluorescence changes.
AuthorsPick U, Finel M
JournalEur J Biochem
PubMed ID6225641
'Chloroplast coupling factor 1 (CF1) contains a high-affinity binding site for 8-anilino-1-napthalene sulphonate (ANS,Kd = 5-6 microM). The binding of ANS to the enzyme is associated with a fluorescence enhancement and a blue-shift in the emission spectrum. ANS only slightly inhibits ATP hydrolysis by CF1. Adenine nucleotides and inorganic phosphate ... More
Assessment of Na+-H+ exchange activity in phagosomal membranes of human neutrophils.
AuthorsGrinstein S, Furuya W
JournalAm J Physiol
PubMed ID2831725
'To assess the presence of Na+-H+ exchange in internalized membranes, the phagosomal pH was monitored in suspensions of intact human neutrophils by measuring the fluorescence of ingested bacteria (Micrococcus lysodeikticus) prelabeled with a pH-sensitive dye. Uptake of fluoresceinated bacteria was confirmed by flow cytometry and by phase-contrast and electron microscopy. ... More
Tethered polymer-supported planar lipid bilayers for reconstitution of integral membrane proteins: silane-polyethyleneglycol-lipid as a cushion and covalent linker.
AuthorsWagner ML, Tamm LK
JournalBiophys J
PubMed ID10969002
'There is increasing interest in supported membranes as models of biological membranes and as a physiological matrix for studying the structure and function of membrane proteins and receptors. A common problem of protein-lipid bilayers that are directly supported on a hydrophilic substrate is nonphysiological interactions of integral membrane proteins with ... More
The organization of F-actin and microtubules in growth cones exposed to a brain-derived collapsing factor.
AuthorsFan J, Mansfield SG, Redmond T, Gordon-Weeks PR, Raper JA
JournalJ Cell Biol
PubMed ID8491778
'In previous work we characterized a brain derived collapsing factor that induces the collapse of dorsal root ganglion growth cones in culture (Raper and Kapfhammer, 1990). To determine how the growth cone cytoskeleton is rearranged during collapse, we have compared the distributions of F-actin and microtubules in normal and partially ... More
A simple fluorescent method for simultaneous determination of aortic permeability to horseradish peroxidase and bovine serum albumin.
AuthorsHollis TM, Katora ME, Montini J
JournalJ Histochem Cytochem
PubMed ID6798104
'Differences in regional aortic net uptake of bovine serum albumin (BSA) and horseradish peroxidase (HP) have been examined by means of conjugation of these molecules to the fluorescent protein tracers fluorescein isothiocyanate (FITC) and lissamine rhodamine B (RB200). Using male Wistar rats, uptake of FITC-BSA under steady state conditions in ... More
A rapid evaluation of phagocytosis and killing of Candida albicans by CD13+ leukocytes.
AuthorsSaresella M, Roda K, Speciale L, Taramelli D, Mendozzi E, Guerini F, Ferrante P
JournalJ Immunol Methods
PubMed ID9520305
'Flow cytometry can be adopted for routine monitoring of the immune functions of human polymorphonuclear leukocytes (PMNs) in several disease states. We recently developed a rapid and reproducible assay for the evaluation of the phagocytosis and killing of Candida albicans blastospores by human PMNs. Whole blood leukocytes were incubated with ... More
In situ localization of paclitaxel binding structures: Labeling with a paclitaxel fluorescent analogue.
AuthorsBicamumpaka C, Page M
JournalInt J Mol Med
PubMed ID9855683
'Microtubules are a major component of cell cytoskeleton. Microtubules constitute the cellular target of paclitaxel. The interaction of paclitaxel with microtubules causes an increase in tubulin polymerization and microtubules stabilization, leading to a G2/M phase cell cycle arrest and cell death by apoptosis. Three paclitaxel fluorescent analogues were prepared by ... More
Diffusion, patching, and capping of stearoylated dextrans on 3T3 cell plasma membranes.
AuthorsWolf DE, Henkart P, Webb WW
JournalBiochemistry
PubMed ID6157402
'Fluorescence-labeled trinitrophenylated stearoylated dextrans have been used as controllable analogues of cell membrane proteins on model membranes and on a variety of natural cell membranes. This paper reports their behavior on 3T3 mouse fibroblast plasma membranes. Spatial distribution on the membrane was studied by fluorescence microscopy, and molecular mobility was ... More
Reaction of bulk protons with a mitochondrial inner membrane preparation: time-resolved measurements and their analysis.
AuthorsGutman M, Kotlyar AB, Borovok N, Nachliel E
JournalBiochemistry
PubMed ID8384483
'The laser-induced proton pulse technique [Gutman, M. (1986) Methods Enzymol. 127, 522-538] was applied on suspensions of submitochondrial vesicles, and the exchange of protons between the bulk and the mitochondrial membranes was measured in the time-resolved domain with a submicrosecond resolution. The protons were discharged by photoexcitation of pyranine (8-hydroxypyrene-1,3,6-trisulfonate) ... More
Effects of supravital fluorochromes used to analyze the in vivo homing of murine lymphocytes on cellular function.
AuthorsSamlowski WE, Robertson BA, Draper BK, Prystas E, McGregor JR
JournalJ Immunol Methods
PubMed ID1960398
'A number of supravital fluorochromes are available to study lymphocyte homing in vivo. These include fluorescein isothiocyanate (FITC), which binds to cell surface proteins; Hoechst 33342, which binds to AT rich regions of cellular DNA; and the lipid bilayer incorporated dyes PKH-2 and PKH-26. The relative advantages and disadvantages of ... More
Fluorescence energy transfer studies of purified erythrocyte Ca2+-ATPase. Ca2+-regulated activation by oligomerization.
AuthorsKosk-Kosicka D, Bzdega T, Wawrzynow A
JournalJ Biol Chem
PubMed ID2531140
'Fluorescence resonance energy transfer has been used to study oligomerization of the purified erythrocyte Ca2+-ATPase. The energy transfer efficiency has been measured at different enzyme concentrations, from fluorescein 5''-isothiocyanate attached on one enzyme molecule to eosin 5-maleimide or tetramethylrhodamine 5-isothiocyanate attached on another enzyme molecule. The energy transfer efficiency showed ... More
Localization of the thrombin-binding domain on prothrombin fragment 2.
AuthorsLiaw PC, Fredenburgh JC, Stafford AR, Tulinsky A, Austin RC, Weitz JI
JournalJ Biol Chem
PubMed ID9535876
'Co-crystallographic studies have shown that the interaction of human prothrombin fragment 2 (F2) with thrombin involves the formation of salt bridges between the kringle inner loop of F2 and anion-binding exosite II of thrombin. When F2 binds to thrombin, it has been shown to evoke conformational changes at the active ... More
Flow cytometric assessment of allopurinol susceptibility in Leishmania infantum promastigote.
AuthorsKamau SW, Hurtado M, Müller-Doblies UU, Grimm F, Nunez R
JournalCytometry
PubMed ID10918286
'BACKGROUND: Leishmaniasis is a major tropical and subtropical parasitic disease. Sodium stibogluconate, N-methyl -D-glucamine antimoniate, amphotericin B, pentamidine, and ketoconazole are drugs used to treat this disease. Some of these drugs cause severe adverse side effects and treatment failures are common. Allopurinol, a purine analog, has been used to treat ... More
A structural change in the Neurospora plasma membrane [H+]ATPase induced by N-ethylmaleimide.
AuthorsChang A, Slayman CW
JournalJ Biol Chem
PubMed ID2144286
'The reaction of N-ethylmaleimide (NEM) with Cys-532 of the Neurospora plasma membrane [H+]ATPase results in inhibition of ATP hydrolysis which is protected by MgADP (Pardo, J. P., and Slayman, C. W. (1989) J. Biol. Chem. 264, 9373-9379). To examine the conformational state of the ATPase upon NEM modification, we have ... More
Labelling of histone H5 and its interaction with DNA. 2. Cooperative binding of histone H5 to DNA as probed by steady-state fluorescence and diffusion-enhanced energy transfer.
AuthorsLerho M, Favazza M, Houssier C
JournalJ Biomol Struct Dyn
PubMed ID2114122
'The interaction of histone H5 labelled with fluorescein isothiocyanate (FITC) with DNA has been studied by fluorescence titration, and diffusion-enhanced fluorescence energy transfer (DEFET) measurements with Tb(III) lanthanide chelates as donors. Analysis of the binding data by the model of Schwarz and Watanabe (J.Mol.Biol. 163, 467-484 (1983)) yielded a mean ... More
A comparison of the emission efficiency of four common green fluorescence dyes after internalization into cancer cells.
AuthorsHama Y, Urano Y, Koyama Y, Bernardo M, Choyke PL, Kobayashi H
JournalBioconjug Chem
PubMed ID17105220
'In vivo optical imaging to enhance the detection of cancer during endoscopy or surgery requires a targeted fluorescent probe with high emission efficiency and high signal-to-background ratio. One strategy to accurately detect cancers is to have the fluorophore internalize within the cancer cells permitting nonbound fluorophores to be washed away ... More
Identification of fluorescein-5'-isothiocyanate-modification sites in proteins by electrospray-ionization mass spectrometry.
AuthorsSchnaible V, Przybylski M
JournalBioconjug Chem
PubMed ID10502354
'Model peptides and proteins, such as hen eggwhite lysozyme, have been modified with fluorescein-5''-isothiocyanate (FITC) to yield the corresponding fluorescein-thiocarbamoyl (FTC) conjugates (N, N''-disubstituted thiourea and dithiourethane adducts). The extent of FITC incorporation, i.e., number of modified residues, has been identified by direct molecular weight determination using matrix-assisted laser desorption-ionization ... More
Nucleus-specific and temporally restricted localization of proteins in Tetrahymena macronuclei and micronuclei.
AuthorsWhite EM, Allis CD, Goldfarb DS, Srivastva A, Weir JW, Gorovsky MA
JournalJ Cell Biol
PubMed ID2553740
'Labeled nuclear proteins were microinjected into the cytoplasm of Tetrahymena thermophila. Macronuclear H1, calf thymus H1, and the SV40 large T antigen nuclear localization signal linked to BSA accumulated specifically in macronuclei, even if cells were in micronuclear S phase or were nonreplicating. The way in which histone H4 localized ... More
Destabilization of peptide binding and interdomain communication by an E543K mutation in the bovine 70-kDa heat shock cognate protein, a molecular chaperone.
AuthorsHa JH, Hellman U, Johnson ER, Li L, McKay DB, Sousa MC, Takeda S, Wernstedt C, Wilbanks SM
JournalJ Biol Chem
PubMed ID9346924
'We have compared 70-kDa heat shock cognate protein (Hsc70) isolated from bovine brain with recombinant wild type protein and mutant E543K protein (previously studied as wild type in our laboratory). Wild type bovine and recombinant protein differ by posttranslational modification of lysine 561 but interact similarly with a short peptide ... More
Optical detection of ion-channel-induced proton transport in supported phospholipid bilayers.
AuthorsYang TH, Yee CK, Amweg ML, Singh S, Kendall EL, Dattelbaum AM, Shreve AP, Brinker CJ, Parikh AN,
JournalNano Lett
PubMed ID17629349
'The integration of ion-channel transport functions with responses derived from nanostructured and nanoporous silica mesophase materials is demonstrated. Patterned thin-film mesophases consisting of alternating hydrophilic nanoporous regions and hydrophobic nanostructured regions allow for spatially localized proton transport via selective dimerization of gramicidin in lipid bilayers formed on the hydrophilic regions. ... More
Applications and perspectives of multi-parameter flow cytometry to microbial biofuels production processes.
Authorsda Silva TL, Roseiro JC, Reis A,
JournalTrends Biotechnol
PubMed ID22257766
'Conventional microbiology methods used to monitor microbial biofuels production are based on off-line analyses. The analyses are, unfortunately, insufficient for bioprocess optimization. Real time process control strategies, such as flow cytometry (FC), can be used to monitor bioprocess development (at-line) by providing single cell information that improves process model formulation ... More
Association of microtubules and intermediate filaments in chicken gizzard cells as detected by double immunofluorescence.
AuthorsGeiger B, Singer SJ
JournalProc Natl Acad Sci U S A
PubMed ID7001467
'By double indirect immunofluorescence, using guinea pig and rabbit antibodies to tubulin and to desmin, we have simultaneously labeled microtubules and intermediate filaments in cultured chicken embryo gizzard cells. At the resolution of the light microscope there was extensive but not complete superposition of the labeling patterns for the two ... More
Calcium activation of the Ca-ATPase enhances conformational heterogeneity between nucleotide binding and phosphorylation domains.
AuthorsChen B, Squier TC, Bigelow DJ
JournalBiochemistry
PubMed ID15065881
'High-resolution crystal structures obtained in two conformations of the Ca-ATPase suggest that a large-scale rigid-body domain reorientation of approximately 50 degrees involving the nucleotide-binding (N) domain is required to permit the transfer of the gamma-phosphoryl group of ATP to Asp(351) in the phosphorylation (P) domain during coupled calcium transport. However, ... More
Rhodopsin in reconstituted phospholipid vesicles. 2. Rhodopsin-rhodopsin interactions detected by resonance energy transfer.
AuthorsBorochov-Neori H, Fortes PA, Montal M
JournalBiochemistry
PubMed ID6830760
'The interactions between rhodopsin molecules in a micellar detergent solution (octyl glucoside) and in reconstituted phospholipid vesicles were studied in the dark and after bleaching. Resonance energy transfer measurements were used to monitor the proximity between rhodopsin monomers conjugated with a fluorescent donor or a fluorescent acceptor. Reactive sulfhydryl groups ... More
Erratic deposition of agrin during the formation of Xenopus neuromuscular junctions in culture.
AuthorsAnderson MJ, Shi ZQ, Grawel R, Zackson SL
JournalDev Biol
PubMed ID7601300
'In order to disclose the mechanism that regulate synapse development we compared the distributions of agrin, acetylcholine receptors (AChR) and a basal lamina heparan sulfate proteoglycan (HSPG) in sections and cultures prepared from Xenopus laevis and Ambystoma mexicanum embryos. While agrin, AChR and HSPG may accumulate almost synchronously at synapses ... More
Quantitative analysis of Fc gamma receptors on murine spleen cell populations by using dual parameter flow cytometry.
AuthorsTitus JA, Finkelman FD, Stephany DA, Jones JF, Segal DM
JournalJ Immunol
PubMed ID6234350
'The expression of Fc gamma R on subsets of mouse spleen cells was examined by dual parameter flow microfluorometry. B cells were detected by labeling them with antibodies against sIgM, sIgD, sIgG, or I-A; essentially all B cells expressed Fc gamma R. The number of Fc gamma R per cell ... More
A heterodimer of thioredoxin and I(B)2 cooperates with Sec18p (NSF) to promote yeast vacuole inheritance.
AuthorsXu Z, Mayer A, Muller E, Wickner W
JournalJ Cell Biol
PubMed ID9015301
'Early in S phase, the vacuole (lysosome) of Saccharomyces cerevisiae projects a stream of vesicles and membranous tubules into the bud where they fuse and establish the daughter vacuole. This inheritance reaction can be studied in vitro with isolated vacuoles. Rapid and efficient homotypic fusion between salt-washed vacuoles requires the ... More
Superoxide dismutase and catalase conjugated to polyethylene glycol increases endothelial enzyme activity and oxidant resistance.
AuthorsBeckman JS, Minor RL, White CW, Repine JE, Rosen GM, Freeman BA
JournalJ Biol Chem
PubMed ID3129432
'Covalent conjugation of superoxide dismutase and catalase with polyethylene glycol (PEG) increases the circulatory half-lives of these enzymes from less than 10 min to 40 h, reduces immunogenicity, and decreases sensitivity to proteolysis. Because PEG has surface active properties and can induce cell fusion, we hypothesized that PEG conjugation could ... More
Affinity labeling of the active site of the Ca2+-ATPase of sarcoplasmic reticulum.
AuthorsMurphy AJ
JournalBiochim Biophys Acta
PubMed ID2974728
'The inactivation of sarcoplasmic reticulum ATPase by fluorescein isothiocyanate (FITC) was shown to have a hyperbolic dependence on the concentration of FITC. The results were quantitatively accounted for by a model in which the reagent first binds reversibly (Kf = 70 microM) to the ATPase and then reacts irreversibly (kmax ... More
Interaction of maleimidobenzoyl actin with myosin subfragment 1 and tropomyosin-troponin.
AuthorsMiki M, Hozumi T
JournalBiochemistry
PubMed ID1827994
'A chemical modification of G-actin with (m-maleimidobenzoyl)-N-hydroxysuccinimide ester (MBS) impairs actin polymerization [Bettache, N., Bertrand, R., & Kassab, R. (1989) Proc. Natl. Acad. Sci. U.S.A. 86, 6028-6032]. MBS-actin recovers the ability to polymerize when a 2-fold molar excess of phalloidin is added in 30 mM KCl/2 mM MgCl2/20 mM Tris-HCl ... More
Kinetics of polymerization of a fluoresceinated derivative of complement protein C9 by the membrane-bound complex of complement proteins C5b-8.
AuthorsSims PJ, Wiedmer T
JournalBiochemistry
PubMed ID6432040
'The fluorescence self-quenching by energy transfer of FITC-C9, a fluoresceinated derivative of human complement protein C9 [Sims, P.J. (1984) Biochemistry (preceding paper in this issue)], has been used to monitor the kinetics of C9 polymerization induced by the membrane-associated complex of complement proteins C5b-8. Time-based measurements of the fluorescence change ... More
Single nucleotide polymorphism detection by combinatorial fluorescence energy transfer tags and biotinylated dideoxynucleotides.
AuthorsTong AK, Ju J
JournalNucleic Acids Res
PubMed ID11861924
'Combinatorial fluorescence energy transfer (CFET) tags, constructed by exploiting energy transfer and combinatorial synthesis, allow multiple biological targets to be analyzed simultaneously. We here describe a multiplex single nucleotide polymorphism (SNP) assay based on single base extension (SBE) using CFET tags and biotinylated dideoxynucleotides (biotin-ddNTPs). A library of CFET-labeled oligonucleotide ... More
Factors underlying the variability of lipid droplet fluorescence in MA-10 Leydig tumor cells.
AuthorsGocze PM, Freeman DA
JournalCytometry
PubMed ID7835165
'Neutral lipids accumulate in cellular lipid droplets. These droplets vary remarkably in number and amount between cells. In the present studies, the variability in lipid content was quantified by comparing the coefficient of variation of fluorescence histograms of nile red lipid-stained cells to the variability of cell size or cell ... More
Physiological inhibitors of the catalytic subunit of cAMP-dependent protein kinase: effect of MgATP on protein-protein interactions.
AuthorsHerberg FW, Taylor SS
JournalBiochemistry
PubMed ID8268180
'The catalytic (C) subunit of cAMP-dependent protein kinase interacts with two classes of inhibitors. The regulatory (R) subunits, types I and II, associate to form an inactive holoenzyme complex that is activated in response to cAMP. The C-subunit is also inhibited by small heat-stable protein kinase inhibitors (PKI''s). Inhibition by ... More
Fluorescent labels influence phagocytosis of Bordetella pertussis by human neutrophils.
AuthorsWeingart CL, Broitman-Maduro G, Dean G, Newman S, Peppler M, Weiss AA
JournalInfect Immun
PubMed ID10417202
'To explore the role of neutrophil phagocytosis in host defense against Bordetella pertussis, bacteria were labeled extrinsically with fluorescein isothiocyanate (FITC) or genetically with green fluorescent protein (GFP) and incubated with adherent human neutrophils in the presence or absence of heat-inactivated human immune serum. In the absence of antibodies, FITC-labeled ... More
Fluorescent vital stains for complementary labelling of protoplasts from Trichoderma spp.
AuthorsHarman GE, Stasz TE
JournalStain Technol
PubMed ID2464211
'In this study several fluorescent vital stains were evaluated for their ability to provide complementary vital staining of protoplasts of Trichoderma spp. for selection of heterokaryons following protoplast fusion. Tetramethyl rhodamine isothiocyanate and fluorescein isothiocyanate were rejected because they stained only a small proportion of protoplasts. Fluorescein diacetate stained all ... More
Switching on of the proliferation or apoptosis of activated human T lymphocytes by IFN-gamma is correlated with the differential expression of the alpha- and beta-chains of its receptor.
AuthorsNovelli F, Bernabei P, Ozmen L, Rigamonti L, Allione A, Pestka S, Garotta G, Forni G
JournalJ Immunol
PubMed ID8757312
'To find out how physiologically secreted IFN-gamma controls either the proliferation or the apoptosis of human T lymphocytes, the kinetics of expression of the alpha- and beta-chains of its receptor (IFN-gamma R) were sequentially followed on T lymphocytes first activated with PHA and then cultured in the presence of IL-2, ... More
Analysis of vertical fluorescence resonance energy transfer from the surface of a small-diameter sphere.
AuthorsJones GM, Wofsy C, Aurell C, Sklar LA
JournalBiophys J
PubMed ID9876165
'Fluorescence resonance energy transfer (FRET) measurements have been used to analyze fluorophore separations in a number of varying geometries, including small particles and extended surfaces. This study focuses on the geometry created by a donor extended above the surface of a small sphere (radius < R0), where the acceptors are ... More
Energy-transfer measurements on a double fluorescent labeled ribonuclease A.
AuthorsJullien M, Garel JR
JournalBiochemistry
PubMed ID6412748
'Two fluorescent groups have been covalently attached to ribonuclease A: first, the alpha-amino group is labeled upon reaction with fluorescein isothiocyanate, and second, one of the active site histidine residues is modified by N-[[(iodoacetyl)amino]ethyl]-5-naphthylamine-1-sulfonic acid. Among the products of these two successive chemical modifications, a derivative bearing one label on ... More
A comparison of non-radioisotopic hybridization assay methods using fluorescent, chemiluminescent and enzyme labeled synthetic oligodeoxyribonucleotide probes.
AuthorsUrdea MS, Warner BD, Running JA, Stempien M, Clyne J, Horn T
JournalNucleic Acids Res
PubMed ID3387214
'N4-[N-(6-trifluoroacetylamidocaproyl)-2-aminoethyl]-5''-O-dimethoxy trityl -5-methyl-2''-deoxycytidine-3''-N,N-diisopropyl-methylphosphoramidite++ + has been synthesized. This N4-alkylamino deoxycytidine derivative has been incorporated into oligonucleotide probes during chemical DNA synthesis. Subsequent to deprotection and purification, fluorescent (fluorescein, Texas Red and rhodamine), chemiluminescent (isoluminol), and enzyme (horseradish peroxidase, alkaline phosphatase) labels have been specifically incorporated. Detection limits of the labels ... More
Release of hydrophobic molecules from polymer micelles into cell membranes revealed by Forster resonance energy transfer imaging.
AuthorsChen H, Kim S, Li L, Wang S, Park K, Cheng JX,
JournalProc Natl Acad Sci U S A
PubMed ID18445654
'It is generally assumed that polymeric micelles, upon administration into the blood stream, carry drug molecules until they are taken up into cells followed by intracellular release. The current work revisits this conventional wisdom. The study using dual-labeled micelles containing fluorescently labeled copolymers and hydrophobic fluorescent probes entrapped in the ... More
Gateable nanofluidic interconnects for multilayered microfluidic separation systems.
'The extension of microfluidic devices to include three-dimensional fluidic networks allows complex fluidic and chemical manipulations but requires innovative methods to interface fluidic layers. Externally controllable interconnects, employing nuclear track-etched polycarbonate membranes containing nanometer-diameter capillaries, are described that produce hybrid three-dimensional fluidic architectures. Controllable nanofluidic transfer is achieved by controlling ... More
Calcium transport by sarcoplasmic reticulum Ca(2+)-ATPase. Role of the A domain and its C-terminal link with the transmembrane region.
AuthorsMöller JV, Lenoir G, Marchand C, Montigny C, le Maire M, Toyoshima C, Juul BS, Champeil P
JournalJ Biol Chem
PubMed ID12138099
'After treatment of sarcoplasmic reticulum Ca(2+)-ATPase with proteinase K (PK) in the presence of Ca(2+) and a protecting non-phosphorylated ligand (e.g. adenosine 5''-(beta,gamma-methylenetriphosphate), we were able to prepare in high yield an ATPase species that only differs from intact ATPase because of excision of the MAATE(243) sequence from the loop ... More
Physical association between MHC class I and class II molecules detected on the cell surface by flow cytometric energy transfer.
AuthorsSzöllösi J, Damjanovich S, Balàzs M, Nagy P, Trón L, Fulwyler MJ, Brodsky FM
JournalJ Immunol
PubMed ID2732468
'The physical association of HLA class I and class II Ag in the membranes of PGF and JY lymphoblastoid cell lines was studied using flow cytometric energy transfer. This technique measures the proximity of cell surface molecules in the nm range and provides a distribution histogram of the average proximity ... More
A high-throughput continuous sample introduction interface for microfluidic chip-based capillary electrophoresis systems.
AuthorsFang Q, Xu GM, Fang ZL
JournalAnal Chem
PubMed ID11922288
'The development of efficient sample introduction and pretreatment systems for microfluidic chip-based analytical systems is important for their application to real-life samples. In this work, world-to-chip interfacing was achieved by a novel flow-through sampling reservoir featuring a guided overflow design. The flow-through reservoir was fabricated on a 30 x 60 ... More
Rotational dynamics and protein-protein interactions in the Ca-ATPase mechanism.
AuthorsSquier TC, Hughes SE, Thomas DD
JournalJ Biol Chem
PubMed ID2837478
'We have varied the degree of protein-protein interactions among Ca-ATPase polypeptide chains in sarcoplasmic reticulum using the cleavable homobifunctional cross-linker dithiobissuccinimidyl propionate and have measured both the rotational mobility and calcium-dependent ATPase activity of the Ca-ATPase in order to assess 1) the nature of the microsecond rotational motion measured by ... More
Studies of protein--DNA interactions by capillary electrophoresis/laser-induced fluorescence polarization.
AuthorsWan QH, Le XC
JournalAnal Chem
PubMed ID11101235
'Protein-DNA interactions were studied on the basis of capillary electrophoretic separation of bound from free fluorescent probe followed by on-line detection with laser-induced fluorescence polarization. Changes in electrophoretic mobility and fluorescence anisotropy upon complex formation were monitored for the determination of binding affinity and stoichiometry. The method was applied to ... More
Fluorescence energy transfer between cobra alpha-toxin molecules bound to the acetylcholine receptor.
AuthorsJohnson DA, Voet JG, Taylor P
JournalJ Biol Chem
PubMed ID6715368
'An approach was developed with steady state fluorescence energy transfer measurements to examine the spatial relationship between the two alpha-toxins bound to the acetylcholine receptor. By taking advantage of the slow dissociation rates of alpha-toxins (Naja naja siamensis 3) from the receptor and of the equal probability with which alpha-toxins ... More
Kinetic aspects of the aggregation of Clostridium perfringens theta-toxin on erythrocyte membranes. A fluorescence energy transfer study.
AuthorsHarris RW, Sims PJ, Tweten RK
JournalJ Biol Chem
PubMed ID2016307
'Fluorescence resonance energy transfer was used to monitor aggregation kinetics of the "thiol-activated" cytolysin (perfringolysin O (PFO) or theta-toxin) of Clostridium perfringens on erythrocyte membranes. PFO was labeled with the isothiocyanate derivatives of either fluorescein or tetramethylrhodamine. No detectable change in the hemolytic activity of PFO was detected after modification ... More
Rotational and conformational dynamics of Escherichia coli ribosomal protein L7/L12.
'Fluorescence methods were utilized to study dynamic aspects of the 24 kDa dimeric Escherichia coli ribosomal protein L7/L12. Oligonucleotide site-directed mutagenesis was used to introduce cysteine residues at specific locations along the peptide chain, in both the C-terminal and N-terminal domains, and various sulfhydryl reactive fluorescence probes (iodoacetamido) fluorescein, IAEDANS, ... More
Reversion from transformed to normal phenotype by inhibition of protein synthesis in rat kidney cells infected with a temperature-sensitive mutant of Rous sarcoma virus.
AuthorsAsh JF, Vogt PK, Singer SJ
JournalProc Natl Acad Sci U S A
PubMed ID185620
'By the use of a rat kidney cell line infected with a temperature-sensitive Rous sarcoma virus, we have shown that, at permissive temperatures where the cells are transformed, concanavalin A induces a clustering of its cell membrane receptors into patches, and the intracellular smooth muscle myosin-like protein is in a ... More
Binding and dissociation of cytochrome c to and from membranes containing acidic phospholipids.
AuthorsSubramanian M, Jutila A, Kinnunen PK
JournalBiochemistry
PubMed ID9477968
'Membrane association and detachment of cytochrome c (cyt c) in millisecond to second time domain were investigated by stopped-flow fluorescence spectroscopy monitoring the efficiency of energy transfer from a pyrene-fatty acid containing phospholipid derivative, 1-palmitoyl-2-[10-(pyren-1-yl)-decanoyl]-sn-glycero-3-phosphoglyce rol (PPDPG, mole fraction X = 0.01) to the heme of the cyt c. Large ... More
Microcontact printing of DNA molecules.
AuthorsLange SA, Benes V, Kern DP, Hörber JK, Bernard A
JournalAnal Chem
PubMed ID15018562
'The controlled placement of DNA molecules onto solid surfaces is the first step in the fabrication of DNA arrays. The sequential deposition of tiny drops containing the probe DNA fragments using arrays of spotting needles or ink jet nozzles has become a standard. However, a caveat of liquid spotting is ... More
Kinetics of multivalent antigen DNP-BSA binding to IgE-Fc epsilon RI in relationship to the stimulated tyrosine phosphorylation of Fc epsilon RI.
AuthorsXu K, Goldstein B, Holowka D, Baird B
JournalJ Immunol
PubMed ID9531278
'Multivalent DNP-BSA is commonly used to cross-link anti-DNP IgE bound to Fc epsilon RI to stimulate cellular responses, although key features of the binding process are unknown. Fluorescence quenching can be used to study the kinetics of DNP-BSA binding to FITC-IgE. We observe that DNP-BSA binds more slowly to IgE ... More
Role of endocardial endothelial cells in the turnover of hyaluronan in Atlantic cod (Gadus morhua).
AuthorsSorensen KK, Dahl LB, Smedsrod B
JournalCell Tissue Res
PubMed ID9377630
'The fate of the major connective tissue polysaccharide hyaluronan, as it appears after release from the matrix, was studied in the Atlantic cod by use of subcutaneous administration of hyaluronan conjugated with fluorescein isothiocyanate (FITC) and labelled with 125I. After administration, the ligand was transported to the heart, which contained ... More
Low affinity interactions of GDPbetaS and ribose- or phosphoryl-substituted GTP analogues with the heterotrimeric G protein, transducin.
'We have examined the effects of three commonly used classes of guanine nucleotide analogues on the retinal G protein, transducin (Gt), and found them to be quite different from those that might be expected from results with other GTP-binding proteins. The most surprising results were with guanosine 5''-O-(2-thiodiphosphate) (GDPbetaS); rather ... More
Lateral diffusion of surface immunoglobulin, Thy-1 antigen, and a lipid probe in lymphocyte plasma membranes.
'Fluorescence photobleaching recovery was used to measure the lateral diffusion coefficient and mobile fraction of surface immunoglobulin (sIg), Thy-1 antigen, and a lipid probe in the plasma membrane of mouse lymphocytes. The lipid probe (3,3''-dioctadecylindocarbocyanine) had a mean (+/-SD) diffusion coefficient of (1.7 +/- 0.3) x 10(-8) cm(2)/sec, with essentially ... More
Determination of liposome size distribution by flow cytometry.
AuthorsVorauer-Uhl K, Wagner A, Borth N, Katinger H
JournalCytometry
PubMed ID10679735
'BACKGROUND: An essential parameter that describes the quality of liposome suspensions is the mean size, respectively the size distribution. Currently several analytical methods including laser light scattering techniques (LLST) are being employed. METHODS: Here we present an alternative technique using flow cytometry (FCM) to characterize uni- and polydisperse suspensions. As ... More
Distribution and function of nitric oxide-containing nerves in canine corpus cavernosum and spongiosum.
AuthorsHedlund P, Larsson B, Alm P, Andersson KE
JournalActa Physiol Scand
PubMed ID8719264
'Specimens of penile erectile tissue from the corpus cavernosum (CC) and corpus spongiosum (CS) of beagle dogs were investigated with reference to morphological and functional aspects of the nitric oxide (NO) system. Using immunohistochemistry, the smooth muscle bundles of the CC were found to contain a rich innervation, as seen ... More
The active site structure of Na+- and K+-stimulated ATPase. Location of a specific fluorescein isothiocyanate reactive site.
'Fluorescein 5''-isothiocyanate (FITC) has been shown to specifically inactivate the Na+- and K+-stimulated adenosine triphosphatase ((Na,K)-ATPase) at low concentrations (Karlish, S. J. D. (1979) Na+,K+ATPase Structure and Kinetics 115-128). The site of modification of purified dog kidney (Na,K)-ATPase by FITC has been investigated by enzymatic cleavage and fluorescence resonance energy ... More
Quantification of effector/target conjugation involving natural killer (NK) or lymphokine activated killer (LAK) cells by two-color flow cytometry.
'Precise estimates of the frequency of NK- and LAK-target conjugates were obtained by two-color flow cytometry using hydroethidine and calcein as intracellular labels for target cells and effector cells, respectively. These two dyes can easily be used with a standard single-laser flow cytometer with excellent signal separation and dye retention. ... More
Labeling of human erythrocyte membranes with eosin probes used for protein diffusion measurements: inhibition of anion transport and photo-oxidative inactivation of acetylcholinesterase.
AuthorsNigg E, Kessler M, Cherry RJ
JournalBiochim Biophys Acta
PubMed ID215229
'The binding of eosin-isothiocyanate (eosin-NCS) and iodoacetamido-eosin (IA-eosin) to band 3 proteins in the membrane of human erythrocytes is characterized by studying the effect of these probes on the anion transport system. Although the unbrominated fluorescein precursors do not affect anion transport, both eosin labels are strong inhibitors of sulphate ... More
Interactions between Eph kinases and ephrins provide a mechanism to support platelet aggregation once cell-to-cell contact has occurred.
AuthorsPrevost N, Woulfe D, Tanaka T, Brass LF,
JournalProc Natl Acad Sci U S A
PubMed ID12084815
'Eph kinases are receptor tyrosine kinases whose ligands, the ephrins, are also expressed on the surface of cells. Interactions between Eph kinases and ephrins on adjacent cells play a central role in neuronal patterning and vasculogenesis. Here we examine the expression of ephrins and Eph kinases on human blood platelets ... More
Evidence for allosteric linkage between exosites 1 and 2 of thrombin.
AuthorsFredenburgh JC, Stafford AR, Weitz JI
JournalJ Biol Chem
PubMed ID9325262
'Investigations to date have demonstrated that ligand binding to exosites 1 or 2 on thrombin produces conformational changes at the active site. In this study, we directly compared the effect of ligand binding to exosites 1 and 2 on the structure and function of the active site of thrombin and ... More