Benzodiazepine binding studies on living cells: application of small ligands for fluorescence correlation spectroscopy.
AuthorsHegener O, Jordan R, Häberlein H
JournalBiol Chem
PubMed ID12530545
'We demonstrate the applicability of fluorescence correlation spectroscopy (FCS) for receptor binding studies using low molecular weight ligands on the membranes of living nerve cells. The binding of the benzodiazepine Ro 7-1986/602 (N-des-diethyl-fluorazepam), labeled with the fluorophore Alexa 532, to the benzodiazepine receptor was analyzed quantitatively at the membrane of ... More
Lateral mobility and specific binding to GABA(A) receptors on hippocampal neurons monitored by fluorescence correlation spectroscopy.
AuthorsMeissner O, Häberlein H
JournalBiochemistry
PubMed ID12578381
'The binding behavior of a fluorescently labeled muscimol derivative to the GABA(A) receptor was analyzed at rat hippocampal neurons by fluorescence correlation spectroscopy. After muscimol had been labeled with the fluorophore Alexa Fluor 532, specific binding constants for binding of the dye-labeled ligand (Mu-Alexa) to the GABA(A) receptor were determined. ... More
Dynamics of beta2-adrenergic receptor-ligand complexes on living cells.
AuthorsHegener O, Prenner L, Runkel F, Baader SL, Kappler J, Häberlein H
JournalBiochemistry
PubMed ID15147203
'The agonist-induced dynamic regulation of the beta(2)-adrenergic receptor (beta(2)-AR) on living cells was examined by means of fluorescence correlation spectroscopy (FCS) using a fluorescence-labeled arterenol derivative (Alexa-NA) in hippocampal neurons and in alveolar epithelial type II cell line A549. Alexa-NA specifically bound to the beta(2)-AR of neurons with a K(D) ... More
Quantitative screening of single copies of human papilloma viral DNA without amplification.
AuthorsLi J, Lee JY, Yeung ES
JournalAnal Chem
PubMed ID16970325
'We describe a novel quantitative viral screening method based on single-molecule detection that does not require amplification. DNA of human papilloma virus (HPV), the major etiological agent of cervical cancer, served as the screening target in this study. Eight 100-nucleotide single-stranded DNA probes were designed complementary to the E6-E7 gene ... More
'Nitric oxide (NO) signaling is inextricably linked to both its physical and chemical properties. Due to its preferentially hydrophobic solubility, NO molecules tend to partition from the aqueous milieu into biological membranes. We hypothesized that plasma membrane ordering provided by cholesterol further couples the physics of NO diffusion with cellular ... More
Detection of allergen-specific IgE on microarrays by use of signal amplification techniques.
AuthorsBacarese-Hamilton T, Mezzasoma L, Ingham C, Ardizzoni A, Rossi R, Bistoni F, Crisanti A
JournalClin Chem
PubMed ID12142398
A novel mechanism for the formation of actin-filament bundles by a nonprocessive formin.
AuthorsMichelot A, Derivery E, Paterski-Boujemaa R, Guérin C, Huang S, Parcy F, Staiger CJ, Blanchoin L,
JournalCurr Biol
PubMed ID17027489
Actin-filament bundles (or cables) have a structural role during cell division and morphogenesis, but also serve as important
Single molecule imaging of protein molecules in nanopores.
AuthorsMa C, Yeung ES,
JournalAnal Chem
PubMed ID20000771
The interactions between single protein molecules and nanoporous polycarbonate membranes were investigated at the single molecule level. Entrapment of proteins was shown to be size selective and was dependent on the membrane pore diameter. A pore size that is only slightly larger than the maximum dimension of the proteins was ... More
Fluorescent DNA hybridization probe preparation using amine modification and reactive dye coupling.
AuthorsCox WG, Singer VL,
JournalBiotechniques
PubMed ID14740493
Fluorescent nucleic acid hybridization probes traditionally have been generated by enzymatic incorporation of dye-labeled nucleotides, even though incorporation efficiency is low and variable from dye to dye. Alternatively, 5-(3-aminoallyl)-2'-deoxyuridine 5'-triphosphate (aa-dUTP) is enzymatically incorporated to generate amine-modified DNA, which is then chemically labeled with an amine-reactive fluorescent dye. We optimized ... More
Reversible, allosteric small-molecule inhibitors of regulator of g protein signaling proteins.
AuthorsBlazer LL, Roman DL, Chung A, Larsen MJ, Greedy BM, Husbands SM, Neubig RR,
JournalMol Pharmacol
PubMed ID20571077
Regulators of G protein signaling (RGS) proteins are potent negative modulators of G protein signaling and have been proposed as potential targets for small-molecule inhibitor development. We report a high-throughput time-resolved fluorescence resonance energy transfer screen to identify inhibitors of RGS4 and describe the first reversible small-molecule inhibitors of an ... More
Ultrasensitive native fluorescence detection of proteins with miniaturized polyacrylamide gel electrophoresis by laser side-entry excitation.
AuthorsZhang H, Yeung ES
JournalElectrophoresis
PubMed ID16927346
Direct detection of separated proteins inside polyacrylamide gels has many advantages compared to staining methods. Ultrasensitive native fluorescence detection of proteins with miniaturized 1-D and 2-D PAGE was achieved with laser side-entry excitation. The detection limit for R-phycoerythrin protein spots in 1-D SDS-PAGE with 532 nm excitation was as low ... More
Efficiencies of fluorescence resonance energy transfer and contact-mediated quenching in oligonucleotide probes.
AuthorsMarras SA, Kramer FR, Tyagi S
JournalNucleic Acids Res
PubMed ID12409481
An important consideration in the design of oligonucleotide probes for homogeneous hybridization assays is the efficiency of energy transfer between the fluorophore and quencher used to label the probes. We have determined the efficiency of energy transfer for a large number of combinations of commonly used fluorophores and quenchers. We ... More
Fluorescent dual colour 2D-protein gel electrophoresis for rapid detection of differences in protein pattern with standard image analysis software.
AuthorsVon Eggeling F, Gawriljuk A, Fiedler W, Ernst G, Claussen U, Klose J, Römer I
JournalInt J Mol Med
PubMed ID11562774
The use of two different fluorescent dyes in two-dimensional (2D) polyacrylamide gel electrophoresis was recently described and termed difference gel electrophoresis (DIGE). Thereby differences between protein samples could be accomplished by fluorescently tagging the samples with different dyes as well as co-separation and visualisation in a single gel. We adapted ... More
Fluorescence-aided molecule sorting: analysis of structure and interactions by alternating-laser excitation of single molecules.
AuthorsKapanidis AN, Lee NK, Laurence TA, Doose S, Margeat E, Weiss S
JournalProc Natl Acad Sci U S A
PubMed ID15175430
We use alternating-laser excitation to achieve fluorescence-aided molecule sorting (FAMS) and enable simultaneous analysis of biomolecular structure and interactions at the level of single molecules. This was performed by labeling biomolecules with fluorophores that serve as donor-acceptor pairs for Förster resonance energy transfer, and by using alternating-laser excitation to excite ... More
Two-photon microscopy and spectroscopy based on a compact confocal scanning head.
AuthorsDiaspro A, Chirico G, Federici F, Cannone F, Beretta S, Robello M
JournalJ Biomed Opt
PubMed ID11516320
We have combined a confocal laser scanning head modified for TPE (two-photon excitation) microscopy with some spectroscopic modules to study single molecules and molecular aggregates. The behavior of the TPE microscope unit has been characterized by means of point spread function measurements and of the demonstration of its micropatterning abilities. ... More
Positive cooperation of protoberberine type 2 alkaloids from Corydalis cava on the GABA(A) binding site.
AuthorsHalbsguth C, Meissner O, Häberlein H
JournalPlanta Med
PubMed ID12709895
Protoberberine alkaloids from the rhizomes of Corydalis cava were investigated with regard to their influence on the GABA A receptor using radioreceptor assays. Whereas the protoberberine type 2 alkaloids, isoapocavidine, corydaline, tetrahydropalmatine, scoulerine and isocorypalmine, increased the specific [(3)H]BMC-binding in a range of 21 - 49 %, the protoberberine type ... More
Automated purification and suspension array detection of 16S rRNA from soil and sediment extracts by using tunable surface microparticles.
AuthorsChandler DP, Jarrell AE
JournalAppl Environ Microbiol
PubMed ID15128511
Autonomous, field-deployable molecular detection systems require seamless integration of complex biochemical solutions and physical or mechanical processing steps. In an attempt to simplify the fluidic requirements for integrated biodetection systems, we used tunable surface microparticles both as an rRNA affinity purification resin in a renewable microcolumn sample preparation system and ... More