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
Nuclear localization signals of human and Thermoplasma proteasomal alpha subunits are functional in vitro.
AuthorsNederlof PM, Wang HR, Baumeister W
JournalProc Natl Acad Sci U S A
PubMed ID8618844
'Proteasomes are located both in the nuclei and in the cytoplasm of eukaryotic cells. Active transport of these complexes through the nuclear pores has been proposed to be mediated by nuclear localization signals (NLS), which have been found in several of the alpha-type proteasomal subunits. We have tested three different ... More
Stability of allophycocyanin's quaternary structure.
AuthorsMacColl R
JournalArch Biochem Biophys
PubMed ID6407396
'The dissociation of allophycocyanin trimers to monomers was examined under a variety of conditions. For alkyl ureas and alcohols the dissociation increased as the straight-chain alkyls increased in length. The effect of branching chains was smaller. Tetrapropylammonium chloride was found to be a very effective agent for trimer dissociation when ... More
Absorption spectrum of allophycocyanin isolated from Anabaena cylindrica: variation of the absorption spectrum induced by changes of the physico-chemical environment.
AuthorsMurakami A, Mimuro M, Ohki K, Fujita Y
JournalJ Biochem (Tokyo)
PubMed ID6783641
'The absorption spectrum of allophycocyanin of Anabaena cylindrica was studied. The extinctions of the main absorption bands (650 and 620 nm) varied depending on the protein concentration, ionic strength, and pH. At higher protein concentrations or higher ionic strength, the 650 nm band became stronger and the 620 nm band ... More
Immunofluorescence measurement in a flow cytometer using low-power helium-neon laser excitation.
AuthorsShapiro HM, Glazer AN, Christenson L, Williams JM, Strom TB
JournalCytometry
PubMed ID6363017
'Helium-neon lasers are economical and efficient light sources; their utility in flow cytometry to date has been limited by the lack of fluorescent probes that can be excited at 633 nm. Allophycocyanin (APC), a highly fluorescent phycobiliprotein, can be used as an antibody label and has spectral characteristics suitable for ... More
Nuclear protein import in permeabilized mammalian cells requires soluble cytoplasmic factors.
AuthorsAdam SA, Marr RS, Gerace L
JournalJ Cell Biol
PubMed ID2391365
'We have developed an in vitro system involving digitonin-permeabilized vertebrate cells to study biochemical events in the transport of macromolecules across the nuclear envelope. While treatment of cultured cells with digitonin permeabilizes the plasma membranes to macromolecules, the nuclear envelopes remain structurally intact and nuclei retain the ability to transport ... More
Three-color immunofluorescence analysis of mouse B-lymphocyte subpopulations.
AuthorsParks DR, Hardy RR, Herzenberg LA
JournalCytometry
PubMed ID6370630
'We have modified a fluorescence-activated cell sorter (FACS) to make three independent immunofluorescence measurements on each cell and used this system to study mouse B-lymphocyte subpopulations. An argon-ion laser (emitting at 488 nm) excites fluorescein- and phycoerythrin-labeled reagents, and a tunable dye laser charged with rhodamine 6G (emitting at 615 ... More
Use of phycoerythrin and allophycocyanin for fluorescence resonance energy transfer analyzed by flow cytometry: advantages and limitations.
AuthorsBatard P, Szollosi J, Luescher I, Cerottini J-C, MacDonald R, Romero P
JournalCytometry
PubMed ID12116371
'This study validates the use of phycoerythrin (PE) and allophycocyanin (APC) for fluorescence energy transfer (FRET) analyzed by flow cytometry.'
Chromophore interactions in allophycocyanin.
AuthorsMacColl R, Csatorday K, Berns DS, Traeger E
JournalBiochemistry
PubMed ID6772206
'Allophycocyanin, which is normally isolated as a trimer (alpha 3 beta 3), has now been successfully dissociated into a monomer (alpha beta) with strikingly different spectroscopic properties. In particular, upon dissociation the characteristic 650-nm absorption and 661-nm fluorescence emission bands of the trimer are completely lost and its fluorescence polarization ... More
Fluorescence properties of allophycocyanin and a crosslinked allophycocyanin trimer.
AuthorsYeh SW, Ong LJ, Clark JH, Glazer AN
JournalCytometry
PubMed ID3100257
'Data on the wavelength and temperature dependence of both time-resolved and steady state fluorescence emission are presented for allophycocyanin (AP) and for a crosslinked allophycocyanin trimer (XL-AP) (Ong LJ and Glazer AN: Physiol Veg 23:777-787, 1985). AP dissociates at high dilution and is not stable above 40 degrees C even ... More
Demonstration of B-cell maturation in X-linked immunodeficient mice by simultaneous three-colour immunofluorescence.
AuthorsHardy RR, Hayakawa K, Parks DR, Herzenberg LA
JournalNature
PubMed ID6358897
'CBA/N mice carrying the X-linked immune deficiency gene (xid) have fewer splenic B cells than normal CBA mice and are unresponsive to a certain class of antigens. Studies of B-cell surface-marker expression and immune responsiveness have led to the commonly accepted idea that the B cells in adult xid mice ... More
Phycobilisome composition and possible relationship to reaction centers.
AuthorsKhanna R, Graham JR, Myers J, Gantt E
JournalArch Biochem Biophys
PubMed ID6408989
'The photosynthetic apparatus was studied in Anacystis nidulans wild type and in a spontaneous pigment mutant 85Y which had improved growth in far-red light (greater than 650 nm). Two phycobiliproteins, C-phycocyanin (lambda max 625) and allophycocyanin (lambda max 650), were present in a molar ratio of approximately 3:1 in the ... More
Kinetics of allophycocyanin's trimer-monomer equilibrium.
AuthorsHuang C, Berns DS, MacColl R
JournalBiochemistry
PubMed ID3103679
'Kinetic studies of the dissociation of allophycocyanin trimers to monomers have been performed by using stopped-flow techniques. The dissociation was monitored by two techniques: by light scattering to observe the molecular weight changes directly and by 650-nm absorbance to observe the linkage of quaternary structure to spectra. The light-scattering experiments ... More
Antioxidative properties of pyruvate and protection of the ischemic rat heart during cardioplegia.
AuthorsDobsak P, Courderot-Masuyer C, Zeller M, Vergely C, Laubriet A, Assem M, Eicher JC, Teyssier JR, Wolf JE, Rochette L
JournalJ Cardiovasc Pharmacol
PubMed ID10547080
'Formation of oxygen free radicals during heart transplantation seems to be related to the alterations occurring during ischemia and reperfusion and could explain the short preservation time of donor hearts. The aim of our study was (a) to analyze the protective effects of pyruvate during cold cardioplegia and ischemia/reperfusion sequence, ... More
The use of fluorescent probes in immunochemistry.
AuthorsDavidson RS, Hilchenbach MM
JournalPhotochem Photobiol
PubMed ID2217553
'The limitations and advantages of particular dyes for labelling proteins and other biological materials are discussed. Methods available for conjugating dyes to proteins are outlined. Following a discussion of double labelling methods the use of photoactivatable fluorochromes and time resolved fluorescence methodologies are outlined. The reasons for the photoinstability of ... More
Spectral properties of phycobilisomes and phycobiliproteins from the blue-green alga-nostoc sp.
AuthorsGray BH, Gantt E
JournalPhotochem Photobiol
PubMed ID806082
Phycobiliproteins.
AuthorsGlazer AN
JournalMethods Enzymol
PubMed ID3148835
Coupling of monoclonal antibodies with fluorophores.
AuthorsHaugland RP
JournalMethods Mol Biol
PubMed ID7550684
Fluorescence-activated cell sorting: theory, experimental optimization, and applications in lymphoid cell biology.
AuthorsParks DR, Herzenberg LA
JournalMethods Enzymol
PubMed ID6396481
Comparative biochemistry of photosynthetic light-harvesting systems.
AuthorsGlazer AN
JournalAnnu Rev Biochem
PubMed ID6412621
Light harvesting by phycobilisomes.
AuthorsGlazer AN
JournalAnnu Rev Biophys Biophys Chem
PubMed ID3924069
The use of phycobiliproteins as fluorescent labels in immunoassay.
AuthorsKronick MN
JournalJ Immunol Methods
PubMed ID3528294
Light guides. Directional energy transfer in a photosynthetic antenna.
AuthorsGlazer AN
JournalJ Biol Chem
PubMed ID2491842
Immunoassay techniques with fluorescent phycobiliprotein conjugates.
AuthorsKronick MN, Grossman PD
JournalClin Chem
PubMed ID6883673
We describe immunoassay techniques that take advantage of the novel properties of phycobiliprotein fluorescent dyes. These dyes, which can be isolated from a wide variety of algae, exhibit extremely high absorptivities, high quantum efficiencies, and excitation and emission bands across the visible spectrum. These stable, hydrophilic proteins can easily be ... More
Simple modification of a commercial flow cytometer to triple laser excitation. Simultaneous five-color fluorescence detection.
AuthorsWoronicz JD, Rice GC
JournalJ Immunol Methods
PubMed ID2661695
We describe a simple and inexpensive modification to the Coulter 753 flow cytometer which enables simultaneous triple laser excitation for advanced multi-color analysis and sorting applications. The salient feature of the modification was to split the rear laser operating in the all lines mode for use in pumping the rhodamine ... More
Separation of phycobiliprotein subunits by reverse-phase high-pressure liquid chromatography.
AuthorsSwanson RV, Glazer AN
JournalAnal Biochem
PubMed ID2221378
Baseline separation of subunits of diverse phycobiliproteins was achieved by a reverse-phase HPLC gradient method with a C4 large-pore column and a solvent system consisting of 0.1% trifluoroacetic acid (TFA) in water and 0.1% TFA in 2:1 (v/v) acetonitrile:isopropanol. The procedure was successfully applied to cyanobacterial allophycocyanin and C-phycocyanins, an ... More
Fluorescent tandem phycobiliprotein conjugates. Emission wavelength shifting by energy transfer.
AuthorsGlazer AN, Stryer L
JournalBiophys J
PubMed ID6414547
A fluorescent tandem phycobiliprotein conjugate with a large Stokes shift was prepared by the covalent attachment of phycoerythrin to allophycocyanin. The efficiency of energy transfer from phycoerythrin to allophycocyanin in this disulfide-linked conjugate was 90%. A distinctive feature of this phycocyanin conjugate is the wide separation between the intense absorption ... More
Near-infrared fluorescence lifetime assay for serum glucose based on allophycocyanin-labeled concanavalin A.
AuthorsMcCartney LJ, Pickup JC, Rolinski OJ, Birch DJ
JournalAnal Biochem
PubMed ID11355853
We describe an assay scheme for glucose based on fluorescence resonance energy transfer (FRET) between concanavalin A (con A), labeled with the near-infrared fluorescent protein allophycocyanin (APC) as donor, and dextran labeled with malachite green (MG) as acceptor. Glucose competitively displaces dextran-MG and leads to reduction in FRET, assessed by ... More
Photostability studies of phycobiliprotein fluorescent labels.
AuthorsWhite JC, Stryer L
JournalAnal Biochem
PubMed ID3578804
Photostability studies of four fluorescent phycobiliproteins were conducted to identify stable chromophores for biological labeling applications. Phycobiliprotein photodestruction was linear with the applied laser power and depended on the total number of photons absorbed per molecule. Photodestruction quantum yields phi of 1.1 X 10(-5) for R-phycoerythrin, 6.6 X 10(-6) for ... More
Assessment of fluorochromes for cellular structure and function studies by flow cytometry.
AuthorsPetit JM, Denis-Gay M, Ratinaud MH
JournalBiol Cell
PubMed ID7693118
Because flow cytometry permits the analysis of individual whole cells, one of the key requirements in selecting a probe is its ability to target the site of interest into cells. In addition, dyes must possess ideal properties (ie extinction coefficient, Stoke's shift) rendering them appropriate for this methodology. Other characteristics, ... More
Rare earth cryptates and homogeneous fluoroimmunoassays with human sera.
AuthorsMathis G
JournalClin Chem
PubMed ID8375081
The rare earth cryptates are used as long-lived fluorescent labels. They are formed by inclusion of a europium ion in the intramolecular cavity of a macropolycyclic ligand containing bipyridine groups as light absorbers. The use of fluoride ions in the measuring medium allows a total shielding of the label fluorescence. ... More
A modular detector for flow cytometric multicolor fluorescence measurements.
AuthorsSteinkamp JA, Habbersett RC, Stewart CC
JournalCytometry
PubMed ID3622158
A modular detector for measuring multicolor fluorescence from cells illuminated by single or multiple lasers has been developed for flow cytometers. Motion picture projector, camera, and CCTV/video lenses were evaluated for use in the detector by comparing their physical characteristics, image quality, and light collection efficiencies. A 25-mm focal length ... More
Chromosomal proteins HMG-14 and HMG-17 are released from mitotic chromosomes and imported into the nucleus by active transport.
AuthorsHock R, Scheer U, Bustin M
JournalJ Cell Biol
PubMed ID9852141
The high mobility group 14/17 (HMG-14/-17) proteins form specific complexes with nucleosome core particles and produce distinct footprints on nucleosomal DNA. Therefore, they could be an integral part of the chromatin fiber. Here we show that during the cell cycle these proteins are transiently dissociated from chromatin. They colocalize with ... More
A practical approach to multicolor flow cytometry for immunophenotyping.
AuthorsBaumgarth N, Roederer M
JournalJ Immunol Methods
PubMed ID10986408
Through a series of novel developments in flow cytometry hardware, software, and dye-chemistry it is now possible to simultaneously measure up to 11 distinct fluorescences and two scattered light parameters on each cell. Such advanced multicolor systems have a number of advantages over current two- and three-color flow cytometric measurements. ... More
8 color, 10-parameter flow cytometry to elucidate complex leukocyte heterogeneity.
AuthorsRoederer M, De Rosa S, Gerstein R, Anderson M, Bigos M, Stovel R, Nozaki T, Parks D, Herzenberg L, Herzenberg L
JournalCytometry
PubMed ID9415416
We developed the chemistry, instrumentation, and software technologies needed to measure, simultaneously and independently, eight different fluorescent molecules on individual cells. Conjugation of these fluorochromes to monoclonal antibodies is straightforward; all immunofluorescence staining is accomplished with direct stains only. We built a hybrid flow cytometer with eight fluorescence detectors and ... More
A novel and inexpensive source of allophycocyanin for multicolor flow cytometry.
AuthorsJung TM, Dailey MO
JournalJ Immunol Methods
PubMed ID2502578
Allophycocyanin (APC) belongs to a family of phycobiliproteins that are well suited as fluorescent reagents for flow cytometric analysis, since they have a broad excitation spectrum, a large Stoke's shift and they fluoresce with a high quantum yield. The widespread use of APC has been limited by the availability of ... More
Ly-1 B-cell clones similar to human chronic lymphocytic leukemias routinely develop in older normal mice and young autoimmune (New Zealand Black-related) animals.
Studies presented here demonstrate that individually expanded clones of murine Ly-1 B cells, perhaps analogous to the expanded neoplastic Leu-1 B-cell clones in human chronic lymphocytic leukemias, are universally detectable in young New Zealand Black (NZB)-related autoimmune mice and in senescent normal mice (greater than 18 months old). These clones ... More
The carboxyl terminus of interferon-gamma contains a functional polybasic nuclear localization sequence.
AuthorsSubramaniam PS, Mujtaba MG, Paddy MR, Johnson HM
JournalJ Biol Chem
PubMed ID9867857
Cytokines such as interferon-gamma (IFN-gamma), which utilize the well studied JAK/STAT pathway for nuclear signal transduction, are themselves translocated to the nucleus. The exact mechanism for the nuclear import of IFN-gamma or the functional role of the nuclear translocation of ligand in signal transduction is unknown. We show in this ... More
Allo-7: a new fluorescent tandem dye for use in flow cytometry.
AuthorsBeavis AJ, Pennline KJ
JournalCytometry
PubMed ID8866224
This report describes the development of a novel tandem dye by combining allophycocyanine (APC) and cyanine dye indotricarbocyanine (CY7) to create ALLO-7 for use in flow cytometry. The APC donor fluorophore was excited at 647 nm and, through resonance energy transfer to the CY7 acceptor, produced fluorescence at > 780 ... More
Fluorescent phycobiliprotein conjugates for analyses of cells and molecules.
AuthorsOi VT, Glazer AN, Stryer L
JournalJ Cell Biol
PubMed ID6749865
The synthesis of a novel class of reagents for fluorescence analyses of molecules and cells is reported. These compounds consist of a highly fluorescent phycobiliprotein conjugated to a molecule having biological specificity. Phycoerythrin-immunoglobulin, phycoerythrin-protein A, and phycoerythrin-avidin conjugates were prepared. These conjugates bind specifically to beads containing a covalently attached ... More
Antioxidant properties of indapamide, 5-OH indapamide and hydrochlorothiazide evaluated by oxygen-radical absorbing capacity and electron paramagnetic resonance.
AuthorsVergely C, Walker MK, Zeller M, Rademakers JR, Maupoil V, Schiavi P, Guez D, Rochette L
JournalMol Cell Biochem
PubMed ID9546594
The aim of these experiments was to investigate the radical scavenging properties of three diuretics: indapamide (IND) and its major metabolite, 5-OH indapamide (5-OH IND), compared to a reference diuretic, hydrochlorothiazide (HTZ). Electron Paramagnetic Resonance (EPR) was used to determine the scavenging abilities of these compounds on enzymatically produced superoxide ... More
Phycoerythrin-allophycocyanin: a resonance energy transfer fluorochrome for immunofluorescence.
AuthorsTjioe I, Legerton T, Wegstein J, Herzenberg LA, Roederer M
JournalCytometry
PubMed ID11309805
BACKGROUND: As immunofluorescence experiments become more complex, the demand for new dyes with different properties increases. Fluorescent dyes with large Stoke's shifts that are very bright and have low background binding to cells are especially desirable. We report on the properties of the resonance energy tandems of phycoerythrin and allophycocyanin ... More
Lymphocyte surface thiol levels.
AuthorsSahaf B, Heydari K, Herzenberg LA, Herzenberg LA
JournalProc Natl Acad Sci U S A
PubMed ID12642656
Recent studies have implicated reduced thiols (cysteine -SH) in the function of individual cell surface proteins. Studies presented here demonstrate that the overall level of reduced thiols on cell surface molecules differs on individual subsets of peripheral blood mononuclear cells and that these levels can be manipulated in vitro by ... More