Inter- and intramolecular fluorescence quenching of organic dyes by tryptophan.
AuthorsMarmé N, Knemeyer JP, Sauer M, Wolfrum J
JournalBioconjug Chem
PubMed ID14624626
'Steady-state and time-resolved fluorescence measurements were performed to elucidate the fluorescence quenching of oxazine, rhodamine, carbocyanine, and bora-diaza-indacene dyes by amino acids. Among the natural amino acids, tryptophan exhibits the most pronounced quenching efficiency. Especially, the red-absorbing dyes ATTO 655, ATTO 680, and the oxazine derivative MR 121 are strongly ... More
Two-color labeling of temporally defined protein populations in mammalian cells.
AuthorsBeatty KE, Tirrell DA,
JournalBioorg Med Chem Lett
PubMed ID18774715
'The proteome undergoes complex changes in response to disease, drug treatment, and normal cellular signaling processes. Characterization of such changes requires methods for time-resolved protein identification and imaging. Here, we describe the application of two reactive methionine (Met) analogues, azidohomoalanine (Aha) and homopropargylglycine (Hpg), to label two protein populations in ... More
Nanoparticle-mediated drug delivery to tumor vasculature suppresses metastasis.
AuthorsMurphy EA, Majeti BK, Barnes LA, Makale M, Weis SM, Lutu-Fuga K, Wrasidlo W, Cheresh DA,
JournalProc Natl Acad Sci U S A
PubMed ID18607000
'Integrin alphanubeta3 is found on a subset of tumor blood vessels where it is associated with angiogenesis and malignant tumor growth. We designed an alphanubeta3-targeted nanoparticle (NP) encapsulating the cytotoxic drug doxorubicin (Dox) for targeted drug delivery to the alphanubeta3-expressing tumor vasculature. We observed real-time targeting of this NP to ... More
Trogocytosis-based generation of suppressive NK cells.
'Trogocytosis is a fast uptake of membranes and associated molecules from one cell by another. Trogocytosis between natural killer (NK) cells and tumors is already described, but the functional relevance of NK-tumor targets material exchange is unclear. We investigated whether the immunosuppressive molecule HLA-G that is commonly expressed by tumors ... More
Toll-like receptor 9-dependent activation by DNA-containing immune complexes is mediated by HMGB1 and RAGE.
AuthorsTian J, Avalos AM, Mao SY, Chen B, Senthil K, Wu H, Parroche P, Drabic S, Golenbock D, Sirois C, Hua J, An LL, Audoly L, La Rosa G, Bierhaus A, Naworth P, Marshak-Rothstein A, Crow MK, Fitzgerald KA, Latz E, Kiener PA, Coyle AJ
JournalNat Immunol
PubMed ID17417641
'Increased concentrations of DNA-containing immune complexes in the serum are associated with systemic autoimmune diseases such as lupus. Stimulation of Toll-like receptor 9 (TLR9) by DNA is important in the activation of plasmacytoid dendritic cells and B cells. Here we show that HMGB1, a nuclear DNA-binding protein released from necrotic ... More
Decreased uptake of bodipy-labelled compounds in the presence of the nuclear stain, DRAQ5.
AuthorsSnyder DS, Garon CF
JournalJ Microsc
PubMed ID12950469
We have found the nuclear stain DRAQ5 to decrease the cellular uptake of a series of boron dipyrromethane (bodipy)-labelled compounds. This phenomenon is consistent between Lysotracker Green DND 26, Lysotracker Red DND 99 and bodipy-labelled mycolactone. Although DRAQ5 uptake was not prevented, DRAQ5 was in significant excess in each case. ... More
Solid-phase oligonucleotide synthesis and flow cytometric analysis with microspheres encoded with covalently attached fluorophores.
AuthorsNanthakumar A, Pon RT, Mazumder A, Yu S, Watson A
JournalBioconjug Chem
PubMed ID10725106
A novel combinatorial approach to synthesize oligonucleotides on fluorescently encoded microspheres based on flow sorting and segmental solid-phase synthesis is described. BODIPY dyes were covalently attached to polystyrene (8.8 microm, 55% DVB) microsphere particles to generate four fluorescently encoded sets. 20-mer oligonucleotide sequences can be synthesized on these microspheres with ... More
Influence of fluorophore and linker composition on the pharmacology of fluorescent adenosine A1 receptor ligands.
AuthorsBaker JG, Middleton R, Adams L, May LT, Briddon SJ, Kellam B, Hill SJ,
JournalBr J Pharmacol
PubMed ID20105183
BACKGROUND AND PURPOSE: The introduction of fluorescence-based techniques, and in particular the development of fluorescent ligands, has allowed the study of G protein-coupled receptor pharmacology at the single cell and single molecule level. This study evaluated how the physicochemical nature of the linker and the fluorophore affected the pharmacological properties ... More
New fluorescent adenosine A1-receptor agonists that allow quantification of ligand-receptor interactions in microdomains of single living cells.
AuthorsMiddleton RJ, Briddon SJ, Cordeaux Y, Yates AS, Dale CL, George MW, Baker JG, Hill SJ, Kellam B
JournalJ Med Chem
PubMed ID17249651
Fluorescence spectroscopy is becoming a valuable addition to the array of techniques available for scrutinizing ligand-receptor interactions in biological systems. In particular, scanning confocal microscopy and fluorescence correlation spectroscopy (FCS) allow the noninvasive imaging and quantification of these interactions in single living cells. To address the emerging need for fluorescently ... More
Application of fluorescence correlation spectroscopy to the measurement of agonist binding to a G-protein coupled receptor at the single cell level.
AuthorsBriddon SJ, Middleton RJ, Yates AS, George MW, Kellam B, Hill SJ
JournalFaraday Discuss
PubMed ID14992407
The A1-adenosine receptor (A1-AR) is a member of the G-protein coupled receptor superfamily, which has significant pathophysiological importance in disorders such as heart arrhythmias, asthma and stroke. Here, we have used fluorescence correlation spectroscopy (FCS) to facilitate the study of A1-AR pharmacology at the subcellular level. To this end, we ... More
Antibiotic-labeled probes and microvolume fluorimetry for the rapid detection of bacterial contamination in platelet components: a preliminary report.
AuthorsBrecher ME, Wong EC, Chen SE, Vampola C, Rocco RM
JournalTransfusion
PubMed ID10773051
BACKGROUND: Approximately 1 platelet in 2000 components is bacterially contaminated. Most commonly, contaminating organisms are gram positive skin saprophytes (such as Staphylococcus sp. or Bacillus sp.). A novel approach to the rapid diagnosis of gram positive contamination by the use of a fluorescence-labeled antibiotic probe with affinity for the gram ... More
Fluorescence intensity and lifetime distribution analysis: toward higher accuracy in fluorescence fluctuation spectroscopy.
AuthorsPalo K, Brand L, Eggeling C, Jäger S, Kask P, Gall K
JournalBiophys J
PubMed ID12124251
Fluorescence fluctuation methods such as fluorescence correlation spectroscopy and fluorescence intensity distribution analysis (FIDA) have proven to be versatile tools for studying molecular interactions with single molecule sensitivity. Another well-known fluorescence technique is the measurement of the fluorescence lifetime. Here, we introduce a method that combines the benefits of both ... More
Fluorescence data analysis on gel-based biochips.
AuthorsBarsky V, Perov A, Tokalov S, Chudinov A, Kreindlin E, Sharonov A, Kotova E, Mirzabekov A
JournalJ Biomol Screen
PubMed ID12097187
A series of biochip readers developed for gel-based biochips includes three imaging models and a novel nonimaging biochip scanner. The imaging readers, ranging from a research-grade versatile reader to a simple portable one, use wide-field objectives and 12-bit digital large-coupled device cameras for parallel addressing of multiple array elements. This ... More
Spectroscopic study and evaluation of red-absorbing fluorescent dyes.
AuthorsBuschmann V, Weston KD, Sauer M
JournalBioconjug Chem
PubMed ID12526709
The spectroscopic characteristics (absorption, emission, and fluorescence lifetime) of 13 commercially available red-absorbing fluorescent dyes were studied under a variety of conditions. The dyes included in this study are Alexa647, ATTO655, ATTO680, Bodipy630/650, Cy5, Cy5.5, DiD, DY-630, DY-635, DY-640, DY-650, DY-655, and EVOblue30. The thorough characterization of this class of ... More
Actin cytoskeletal reorganizations and coreceptor-mediated activation of rac during human immunodeficiency virus-induced cell fusion.
AuthorsPontow SE, Heyden NV, Wei S, Ratner L
JournalJ Virol
PubMed ID15194790
The membrane fusion events which initiate human immunodeficiency virus type 1 (HIV-1) infection and promote cytopathic syncytium formation in infected cells commence with the binding of the HIV envelope glycoprotein (Env) to CD4 and an appropriate coreceptor. Here, we show that HIV Env-coreceptor interactions activate Rac-1 GTPase and stimulate the ... More
Bivariate analysis of cellular DNA versus RNA content by laser scanning cytometry using the product of signal subtraction (differential fluorescence) as a separate parameter.
AuthorsSmolewski P, Grabarek J, Kamentsky LA, Darzynkiewicz Z
JournalCytometry
PubMed ID11598949
BACKGROUND: The cytometric methods of bivariate analysis of cellular RNA versus DNA content have limitations. The method based on the use of metachromatic fluorochrome acridine orange (AO) requires rigorous conditions of the equilibrium staining whereas pyronin Y and Hoechst 33342 necessitate the use of an instrument that provides two-laser excitation, ... More
Quantitative analysis of the formation and diffusion of A1-adenosine receptor-antagonist complexes in single living cells.
AuthorsBriddon SJ, Middleton RJ, Cordeaux Y, Flavin FM, Weinstein JA, George MW, Kellam B, Hill SJ
JournalProc Natl Acad Sci U S A
PubMed ID15070776
The A1-adenosine receptor (A1-AR) is a G protein-coupled receptor that mediates many of the physiological effects of adenosine in the brain, heart, kidney, and adipocytes. Currently, ligand interactions with the A1-AR can be quantified on large cell populations only by using radioligand binding. To increase the resolution of these measurements, ... More
Four-color DNA sequencing by synthesis on a chip using photocleavable fluorescent nucleotides.
AuthorsSeo TS, Bai X, Kim DH, Meng Q, Shi S, Ruparel H, Li Z, Turro NJ, Ju J
JournalProc Natl Acad Sci U S A
PubMed ID15829588
We report four-color DNA sequencing by synthesis (SBS) on a chip, using four photocleavable fluorescent nucleotide analogues (dGTP-PC-Bodipy-FL-510, dUTP-PC-R6G, dATP-PC-ROX, and dCTP-PC-Bodipy-650) (PC, photocleavable; Bodipy, 4,4-difluoro-4-bora-3alpha,4alpha-diaza-s-indacene; ROX, 6-carboxy-X-rhodamine; R6G, 6-carboxyrhodamine-6G). Each nucleotide analogue consists of a different fluorophore attached to the 5 position of the pyrimidines and the 7 position ... More