Quantitation of polymerase chain reaction products by capillary electrophoresis using laser fluorescence.
AuthorsButler JM, McCord BR, Jung JM, Wilson MR, Budowle B, Allen RO
JournalJ Chromatogr B Biomed Appl
PubMed ID7820255
In samples where the amount of DNA is limited, the polymerase chain reaction (PCR) can amplify specific regions of the DNA. A quantitative analysis of the PCR product would be desirable to ensure sufficient DNA is available for analysis. In this study, we examine the use of capillary electrophoresis (CE) ... More
Drosophila atypical protein kinase C associates with Bazooka and controls polarity of epithelia and neuroblasts.
AuthorsWodarz A, Ramrath A, Grimm A, Knust E
JournalJ Cell Biol
PubMed ID10995441
'The establishment and maintenance of polarity is of fundamental importance for the function of epithelial and neuronal cells. In Drosophila, the multi-PDZ domain protein Bazooka (Baz) is required for establishment of apico-basal polarity in epithelia and in neuroblasts, the stem cells of the central nervous system. In the latter, Baz ... More
Asbestos increases mammalian AP-endonuclease gene expression, protein levels, and enzyme activity in mesothelial cells.
'Only two DNA repair enzymes, DNA polymerase beta and O6-methylguanine-DNA methyltransferase, have been shown to be inducible in mammalian cells by genotoxic agents. We show here that crocidolite asbestos induces the DNA repair enzyme, apurinic/apyrimidinic (AP)-endonuclease, in isolated mesothelial cells, the progenitor cells of malignant mesothelioma. Asbestos at nontoxic concentrations ... More
Monomethylation of histone H4-lysine 20 is involved in chromosome structure and stability and is essential for mouse development.
AuthorsOda H, Okamoto I, Murphy N, Chu J, Price SM, Shen MM, Torres-Padilla ME, Heard E, Reinberg D,
JournalMol Cell Biol
PubMed ID19223465
'PR-Set7/Set8/KMT5A is the sole enzyme known to catalyze monomethylation of histone H4 lysine 20 (H4K20) and is present only in multicellular organisms that compact a large fraction of their DNA. We found that mouse embryos that are homozygous null mutants for the gene PR-Set7 display early embryonic lethality prior to ... More
Sensitive multicolor fluorescence in situ hybridization using catalyzed reporter deposition (CARD) amplification.
AuthorsSpeel EJ, Ramaekers FC, Hopman AH
JournalJ Histochem Cytochem
PubMed ID9313806
'We describe the simultaneous localization of DNA sequences in cell and chromosome preparations by means of differently fluorochrome-labeled (AMCA, FITC, TRITC) tyramides using the catalyzed reporter deposition (CARD) procedure. For this purpose, repeated as well as single-copy DNA probes were labeled with biotin, digoxigenin, and FITC, hybridized, and visualized with ... More
Evidence that Sry is expressed in pre-Sertoli cells and Sertoli and granulosa cells have a common precursor.
AuthorsAlbrecht KH, Eicher EM
JournalDev Biol
PubMed ID11784049
'The expression of Sry in the undifferentiated, bipotential genital ridges of mammalian XY fetuses initiates testis development and is hypothesized to do so by directing supporting cell precursors to develop as Sertoli cells and not as granulosa cells. To directly test this hypothesis, transgenic mice expressing EGFP under the control ... More
Quantifying single gene copy number by measuring fluorescent probe lengths on combed genomic DNA.
AuthorsHerrick J, Michalet X, Conti C, Schurra C, Bensimon A
JournalProc Natl Acad Sci U S A
PubMed ID10618399
'An approach was developed for the quantification of subtle gains and losses of genomic DNA. The approach relies on a process called molecular combing. Molecular combing consists of the extension and alignment of purified molecules of genomic DNA on a glass coverslip. It has the advantage that a large number ... More
A new method for imaging and 3D reconstruction of mammalian cochlea by fluorescent confocal microscopy.
AuthorsHardie NA, MacDonald G, Rubel EW
JournalBrain Res
PubMed ID15053969
'Traditional methods for anatomical and morphometric studies of cochlear tissues have relied upon either microdissection of the organ of Corti or the generation of serial sections of the cochlea. Such methods are time-consuming, disruptive to three-dimensional relationships and often restrict sampling to very limited numbers of cells. We have found ... More
Ribosomes in the squid giant axon.
AuthorsBleher R, Martin R
JournalNeuroscience
PubMed ID11719007
'Ribosome clusters, referred to as endoaxoplasmic plaques, were documented and quantitatively analyzed in the squid giant axon at the light and electron microscopic levels. The methods included nonspecific high affinity fluorescence staining of RNA by YOYO-1, specific immunofluorescence labeling of ribosomal RNA, electron energy loss spectroscopic mapping of ribosomal phosphorus, ... More
Methylated DNA-binding proteins from Arabidopsis.
AuthorsIto M, Koike A, Koizumi N, Sano H
JournalPlant Physiol
PubMed ID14605234
'The 5-methylcytosines (m5C) play a critical role in epigenetic control, often being recognized by proteins containing a methyl-CpG-binding domain (MBD). Database screening has identified at least 12 putative methyl-CpG-binding proteins from Arabidopsis; we have isolated corresponding cDNAs for seven of them. Despite variation in size and amino acid sequence, all ... More
Anomalous radial migration of single DNA molecules in capillary electrophoresis.
AuthorsZheng J, Yeung ES
JournalAnal Chem
PubMed ID12236367
'We report the unexpected radial migration of DNA molecules in capillary electrophoresis (CE) with applied Poiseuille flow. Such movement can contribute to anomalous migration times, peak dispersion, and size and shape selectivity in CE. When Poiseuille flow is applied from the cathode to the anode, DNA molecules move toward the ... More
Ultrasensitive detection of genetically modified maize DNA by capillary gel electrophoresis with laser-induced fluorescence using different fluorescent intercalating dyes.
AuthorsGarcía-Cañas V, González R, Cifuentes A
JournalJ Agric Food Chem
PubMed ID12137467
'In this work, four different fluorescent intercalating dyes are compared for the ultrasensitive CGE-LIF detection of DNA from transgenic maize in flours. The fluorescent intercalating dyes compared are YOPRO-1, SYBR-Green-I, Ethidium bromide (EthBr), and EnhanCE. For all the four dyes optimum concentrations are established, and efficient separations of DNA fragments ... More
Protein synthesizing units in presynaptic and postsynaptic domains of squid neurons.
AuthorsMartin R, Vaida B, Bleher R, Crispino M, Giuditta A
JournalJ Cell Sci
PubMed ID9763510
'Putative protein synthesizing domains, called plaques, are characterized in the squid giant synapse and axon and in terminals of squid photoreceptor neurons. Plaques are oval-shaped formations of about 1 microm in size, which (1) generate signals that have spectroscopic electron energy loss characteristics of ribosomes, (2) exhibit ribonuclease-sensitive binding of ... More
Protective effect of vitamin C against double-strand breaks in reconstituted chromatin visualized by single-molecule observation.
AuthorsYoshikawa Y, Hizume K, Oda Y, Takeyasu K, Araki S, Yoshikawa K
JournalBiophys J
PubMed ID16284271
'Direct attack to genomic DNA by reactive oxygen species causes various types of lesions, including base modifications and strand breaks. The most significant lesion is considered to be an unrepaired double-strand break that can lead to fatal cell damage. We directly observed double-strand breaks of DNA in reconstituted chromatin stained ... More
Differential effects of deuterium oxide on the fluorescence lifetimes and intensities of dyes with different modes of binding to DNA.
AuthorsSailer BL, Nastasi AJ, Valdez JG, Steinkamp JA, Crissman HA
JournalJ Histochem Cytochem
PubMed ID9016307
'Deuterium oxide (D2O) increases both the fluorescence lifetime and the fluorescence intensity of the intercalating dyes propidium iodide (PI) and ethidium bromide (EB) when bound to nucleic acid structures. We have used spectroscopic analysis coupled with conventional and phase-sensitive flow cytometry to compare the alterations in intensity and lifetime of ... More
1H NMR studies of the bis-intercalation of a homodimeric oxazole yellow dye in DNA oligonucleotides.
AuthorsJohansen F, Jacobsen JP
JournalJ Biomol Struct Dyn
PubMed ID9833661
'We have used one and two dimensional 1H NMR spectroscopy to characterize the binding of a homodimeric oxazole yellow dye, 1,1'-(4,4,8,8-tetramethyl-4,8-diaza-undecamethylene)-bis-4-( 3-methyl-2,3-dihydro-(benzo-1,3-oxazole)-2-methylidene)-quinoliniu m tetraiodide (YOYO), to oligonucleotides containing the (5'-CTAG-3')2 and the (5'-CCGG-3')2 binding sites in either different oligonucleotides or in the same oligonucleotide. YOYO bis-intercalates strongly in all the ... More
Stable fluorescent complexes of double-stranded DNA with bis-intercalating asymmetric cyanine dyes: properties and applications.
AuthorsRye HS, Yue S, Wemmer DE, Quesada MA, Haugland RP, Mathies RA, Glazer AN
JournalNucleic Acids Res
PubMed ID1614866
'The synthesis, proof of structure, and the absorption and fluorescence properties of two new unsymmetrical cyanine dyes, thiazole orange dimer (TOTO; 1,1''-(4,4,7,7-tetramethyl-4,7- diazaundecamethylene)-bis-4-[3-methyl-2,3-dihydro-(benzo-1,3-thiaz ole)-2- methylidene]-quinolinium tetraiodide) and oxazole yellow dimer (YOYO; an analogue of TOTO with a benzo-1,3-oxazole in place of the benzo-1,3-thiazole) are reported. TOTO and YOYO are virtually ... More
Spectroscopic studies of single-stranded DNA ligands and oxazole yellow dyes.
AuthorsAbramo KH, Pitner JB, McGown LB
JournalBiospectroscopy
PubMed ID9547012
'Interactions between short single-stranded DNA ligands and fluorescent DNA indicator dyes were used to investigate binding selectivity of the ligands. Conformational differences among four DNA ligands of different sequence and structure, including two that form a G-quartet and two that do not, were confirmed by circular dichroism spectroscopy. Their interactions ... More
Snapshot blotting: transfer of nucleic acids and nucleoprotein complexes from electrophoresis gels to grids for electron microscopy.
AuthorsJett SD, Bear DG
JournalProc Natl Acad Sci U S A
PubMed ID8041711
'We present a technique, "snapshot blotting," for the electrophoretic transfer of nucleic acids and nucleoprotein complexes in gel electrophoresis bands onto highly stable carbon film-coated grids for imaging by electron microscopy. The method permits structural analysis of macromolecular species that have been resolved by a gel mobility-shift assay. To demonstrate ... More
Selective detection of individual DNA molecules by capillary polymerase chain reaction.
AuthorsLi H, Xue G, Yeung ES
JournalAnal Chem
PubMed ID11321306
'On-line capillary polymerase chain reaction (PCR) coupled with laser-induced fluorescence detection was successfully demonstrated for individual DNA molecules. A single 30-microm-i.d. fused-silica capillary was used both as the reaction vessel and for isolating single molecules. SYBR green I dye was added into the reaction mixture for dynamic fluorescent labeling. Because ... More
X-ray induced DNA double-strand breaks in human sperm.
AuthorsSingh NP, Stephens RE
JournalMutagenesis
PubMed ID9491398
'A methodology for quantifying DNA double-strand breaks in human sperm is described. Sperm from three healthy human donors on three separate days each were irradiated with 12.5, 25, 50 and 100 cGy X-rays. Linear dose-response effects were observed in migrated DNA from sperm nuclei when electrophoresed under neutral conditions. RNase ... More
Structure-function analysis of the protein-binding domains of Mac1p, a copper-dependent transcriptional activator of copper uptake in Saccharomyces cerevisiae.
AuthorsSerpe M, Joshi A, Kosman DJ
JournalJ Biol Chem
PubMed ID10506178
'The Mac1 protein in Saccharomyces cerevisiae is essential for the expression of yeast high affinity copper uptake. A positive transcription factor, Mac1p binds via its N-terminal domain to GCTC elements in the promoters of CTR1 and FRE1, encoding a copper permease and metal reductase, respectively. Mac1p-dependent transcriptional activation is negatively ... More
High-sensitivity capillary electrophoresis of double-stranded DNA fragments using monomeric and dimeric fluorescent intercalating dyes.
AuthorsZhu H, Clark SM, Benson SC, Rye HS, Glazer AN, Mathies RA
JournalAnal Chem
PubMed ID8067520
'Fluorescence-detected capillary electrophoresis separations of phi X174/HaeIII DNA restriction fragments have been performed using monomeric and dimeric intercalating dyes. Replaceable hydroxyethyl cellulose solutions were used as the separation medium. Confocal fluorescence detection was performed following 488-nm laser excitation. The limits of DNA detection for on-column staining with monomeric dyes (ethidium ... More
Stable fluorescent dye-DNA complexes in high sensitivity detection of protein-DNA interactions. Application to heat shock transcription factor.
AuthorsRye HS, Drees BL, Nelson HC, Glazer AN
JournalJ Biol Chem
PubMed ID8227088
'The gel mobility-shift assay is an important tool for the study of protein-nucleic acid interactions. High detection sensitivity is typically attained by radioisotopic labeling of the target nucleic acid fragments. A novel fluorescence methodology offers significant advantages over this conventional approach. Ethidium, thiazole orange, and oxazole yellow homodimers form stable, ... More
Green-light transilluminator for the detection without photodamage of proteins and DNA labeled with different fluorescent dyes.
AuthorsAlba FJ, Bermúdez A, Daban JR
JournalElectrophoresis
PubMed ID11258745
'The excitation spectra of Nile red and SYPRO red, two currently used dyes for the fluorescent staining of protein bands in sodium dodecyl sulfate (SDS)-polyacrylamide gels, show an excitation peak in the UV region and another in the visible region (maximum at about 550 nm). Ethidium bromide and other intercalating ... More
Sharp DNA bends as landmarks of protein-binding sites on straightened DNA.
AuthorsYokota H, Fung K, Trask BJ, van den Engh G, Sarikaya M, Aebersold R
JournalAnal Chem
PubMed ID10330902
'We have developed a fluorescence-based method for mapping single or multiple protein-binding sites on straightened, large-size DNA molecules (> 5 kbp). In the described method, protein-DNA complexes were straightened and immobilized on a flat surface using surface tension. A fraction of the immobilized complexes displayed a sharp DNA bend with ... More
Polarization of scatter and fluorescence signals in flow cytometry.
AuthorsAsbury CL, Uy JL, van den Engh G
JournalCytometry
PubMed ID10805928
'BACKGROUND: The pulses of light scatter and fluorescence measured in flow cytometers exhibit varying degrees of polarization. Flow cytometers are heterogeneously sensitive to this polarization, depending on the light source(s), the optical layout, and the types of mirrors and filters used. Therefore, fluorescence polarization can affect apparent intensity ratios between ... More
Spectral measurements of intercalated PCR-amplified short tandem repeat alleles.
AuthorsMarino MA, Devaney JM, Davis PA, Smith JK, Girard JE
JournalAnal Chem
PubMed ID9823710
'Short tandem repeat (STR) alleles are popular for use as forensic markers due to their highly polymorphic nature. Commonly they are separated by gel electrophoresis and visualized using intercalation dyes. The purpose of this study was to determine the changes in absorbance and fluorescence of DNA-intercalation dye complexes as a ... More
Nano-scale imaging of chromosomes and DNA by scanning near-field optical/atomic force microscopy.
AuthorsYoshino T, Sugiyama S, Hagiwara S, Fukushi D, Shichiri M, Nakao H, Kim JM, Hirose T, Muramatsu H, Ohtani T
JournalUltramicroscopy
PubMed ID12801660
'Nano-scale structures of the YOYO-1-stained barley chromosomes and lambda-phage DNA were investigated by scanning near-field optical/atomic force microscopy (SNOM/AFM). This technique enabled precise analysis of fluorescence structural images in relation to the morphology of the biomaterials. The results suggested that the fluorescence intensity does not always correspond to topographic height ... More
Role of endocytosis in the transfection of L929 fibroblasts by polyethylenimine/DNA complexes.
AuthorsRémy-Kristensen A, Clamme JP, Vuilleumier C, Kuhry JG, Mély Y
JournalBiochim Biophys Acta
PubMed ID11513802
'Polyethylenimine (PEI) is one of the most efficient nonviral vectors for gene therapy. The aim of this study was to investigate the role of endocytosis in the transfection of synchronized L929 fibroblasts by PEI/DNA complexes. This was performed by confocal microscopy and flow cytometry, using the endocytosis marker FM4-64 and ... More
Dynamics of individual flexible polymers in a shear flow.
AuthorsLeDuc P, Haber C, Bao G, Wirtz D
JournalNature
PubMed ID10376595
'Polymer dynamics are of central importance in materials science, mechanical engineering, biology and medicine. The dynamics of macromolecular solutions and melts in shear flow are typically studied using bulk experimental methods such as light and neutron scattering and birefringence. But the effect of shear on the conformation and dynamics of ... More
Comet assay with nuclear extract incubation.
AuthorsWang AS, Ramanathan B, Chien YH, Goparaju CM, Jan KY
JournalAnal Biochem
PubMed ID15649377
'Alkaline comet assay is a simple sensitive method for detecting DNA strand breaks. However, at the time of cell lysis, only a fraction of the entire DNA damage appears as DNA strand breaks, while some DNA strand breaks may have been rejoined and some DNA lesions may still remain unexcised. ... More
Quantitative RNA-polymerase chain reaction-DNA analysis by capillary electrophoresis and laser-induced fluorescence.
AuthorsFasco MJ, Treanor CP, Spivack S, Figge HL, Kaminsky LS
JournalAnal Biochem
PubMed ID7710061
'Quantitative RNA-polymerase chain reaction (RNA-PCR) is an extremely powerful analytical tool owing to its specificity and high level of sensitivity. Quantitative RNA-PCR is, however, highly labor intensive. No analytical method currently exists that can accurately and rapidly quantitate the small quantities of DNA in RNA-PCR reaction mixtures. We have developed ... More
Single molecule analysis of DNA replication.
AuthorsHerrick J, Bensimon A
JournalBiochimie
PubMed ID10572299
'We describe here a novel approach for the study of DNA replication. The approach is based on a process called molecular combing and allows for the genome wide analysis of the spatial and temporal organization of replication units and replication origins in a sample of genomic DNA. Molecular combing is ... More
TOTO and YOYO: new very bright fluorochromes for DNA content analyses by flow cytometry.
AuthorsHirons GT, Fawcett JJ, Crissman HA
JournalCytometry
PubMed ID7513274
'Flow cytometric (FCM) studies were performed on nuclei, ethanol-fixed CHO cells, and isolated human GM130 chromosomes stained with two new cyanine dyes, TOTO and YOYO. These fluorochromes, which are dimers of thiazole orange and oxazole yellow, respectively, have high quantum efficiencies and exhibit specificities for both DNA and RNA. Bound ... More
Design and formulation of polyplexes based on pluronic-polyethyleneimine conjugates for gene transfer.
'Previously, we reported the evaluation of several polyplex-based gene delivery systems with respect to their effectiveness, toxicity, and cell type dependence in vitro. One system, P123-g-PEI(2K), a cationic graft block copolymer, is of particular interest as it has been demonstrated to successfully deliver genetic material to murine liver following systemic ... More
Whole-genome shotgun optical mapping of Rhodobacter sphaeroides strain 2.4.1 and its use for whole-genome shotgun sequence assembly.
AuthorsZhou S, Kvikstad E, Kile A, Severin J, Forrest D, Runnheim R, Churas C, Hickman JW, Mackenzie C, Choudhary M, Donohue T, Kaplan S, Schwartz DC
JournalGenome Res
PubMed ID12952882
'Rhodobacter sphaeroides 2.4.1 is a facultative photoheterotrophic bacterium with tremendous metabolic diversity, which has significantly contributed to our understanding of the molecular genetics of photosynthesis, photoheterotrophy, nitrogen fixation, hydrogen metabolism, carbon dioxide fixation, taxis, and tetrapyrrole biosynthesis. To further understand this remarkable bacterium, and to accelerate an ongoing sequencing project, ... More
Gene transfer by cationic surfactants is essentially limited by the trapping of the surfactant/DNA complexes onto the cell membrane: a fluorescence investigation.
AuthorsClamme JP, Bernacchi S, Vuilleumier C, Duportail G, Mély Y
JournalBiochim Biophys Acta
PubMed ID11030593
'The interaction between complexes of plasmid DNA with cetyltrimethylammonium bromide (CTAB) and L929 fibroblasts was first examined using confocal microscopy. The complexes labeled with the DNA intercalator, YOYO-1, were found to be trapped onto the external face of the plasma membrane; a feature that may constitute a major limiting step ... More
The two nuclei of Giardia each have complete copies of the genome and are partitioned equationally at cytokinesis.
AuthorsYu LZ, Birky CW, Adam RD
JournalEukaryot Cell
PubMed ID12455954
'Giardia lamblia is medically important as a cause of diarrhea and malabsorption throughout the world and is thought to be one of the earliest-branching eukaryotes on a phylogenetic tree. Nevertheless, the mechanisms of inheritance are largely unknown. The trophozoites of Giardia and other diplomonads are interesting in their possession of ... More
Groove-binding unsymmetrical cyanine dyes for staining of DNA: syntheses and characterization of the DNA-binding.
AuthorsKarlsson HJ, Eriksson M, Perzon E, Akerman B, Lincoln P, Westman G
JournalNucleic Acids Res
PubMed ID14576310
'Two new crescent-shaped unsymmetrical cyanine dyes have been synthesised and their interactions with DNA have been investigated by different spectroscopic methods. These dyes are analogues to a minor groove binding unsymmetrical cyanine dye, BEBO, recently reported by us. In this dye, the structure of the known intercalating cyanine dye BO ... More
Interaction of oxazole yellow dyes with DNA studied with hybrid optical tweezers and fluorescence microscopy.
AuthorsMurade CU, Subramaniam V, Otto C, Bennink ML,
JournalBiophys J
PubMed ID19651041
'We have integrated single molecule fluorescence microscopy imaging into an optical tweezers set-up and studied the force extension behavior of individual DNA molecules in the presence of various YOYO-1 and YO-PRO-1 concentrations. The fluorescence modality was used to record fluorescent images during the stretching and relaxation cycle. Force extension curves ... More
Modifications of alkaline microgel electrophoresis for sensitive detection of DNA damage.
AuthorsSingh NP, Stephens RE, Schneider EL
JournalInt J Radiat Biol
PubMed ID8027608
'The alkaline microgel electrophoresis technique was modified to achieve a substantial increase in sensitivity for the detection of radiation-induced DNA damage in human lymphocytes. This increased sensitivity was achieved through: (1) the addition of free radical scavengers to the electrophoresis solution to reduce DNA damage generated during alkaline unwinding and ... More
Organized arrays of individual DNA molecules tethered to supported lipid bilayers.
AuthorsGranéli A, Yeykal CC, Prasad TK, Greene EC
JournalLangmuir
PubMed ID16378434
'An unappreciated aspect of many single-molecule techniques is the need for an inert surface to which individual molecules can be anchored without compromising their biological integrity. Here, we present new methods for tethering large DNA molecules to the surface of a microfluidic sample chamber that has been rendered inert by ... More
Observation by fluorescence microscopy of transcription on single combed DNA.
AuthorsGueroui Z, Place C, Freyssingeas E, Berge B
JournalProc Natl Acad Sci U S A
PubMed ID11983896
'Molecular combing is a powerful procedure for aligning a large array of DNA molecules onto a surface. This technique usually leads to an overstretching of about 150% of the molecules'' contour length. By changing the magnitude of capillary forces during the combing process, we were able to reduce the relative ... More
Fluorescent DNA nanotags: supramolecular fluorescent labels based on intercalating dye arrays assembled on nanostructured DNA templates.
AuthorsBenvin AL, Creeger Y, Fisher GW, Ballou B, Waggoner AS, Armitage BA
JournalJ Am Chem Soc
PubMed ID17256855
'Fluorescence detection and imaging are vital technologies in the life sciences and clinical diagnostics. The key to obtaining high-resolution images and sensitive detection is to use fluorescent molecules or particles that absorb and emit visible light with high efficiency. We have synthesized supramolecular complexes consisting of a branched DNA template ... More
Spatial organization of large-scale chromatin domains in the nucleus: a magnified view of single chromosome territories.
AuthorsFerreira J, Paolella G, Ramos C, Lamond AI
JournalJ Cell Biol
PubMed ID9412456
'We have analyzed the spatial organization of large scale chromatin domains in chinese hamster fibroblast, human lymphoid (IM-9), and marsupial kidney epithelial (PtK) cells by labeling DNA at defined stages of S phase via pulsed incorporation of halogenated deoxynucleosides. Most, if not all, chromosomes contribute multiple chromatin domains to both ... More
High-speed screening of polymerase chain reaction products by capillary electrophoresis.
'In an effort to develop capillary electrophoresis (CE) for high-throughput polymerase chain reaction (PCR) molecular diagnostics, a method was developed to rapidly screen small PCR products of similar molecular weights. The assay of interest required the separation of two PCR products (375 and 400 bp) in an assay of TGF-beta ... More
Opposing actions of inositol 1,4,5-trisphosphate and ryanodine receptors on nuclear factor of activated T-cells regulation in smooth muscle.
AuthorsGomez MF, Stevenson AS, Bonev AD, Hill-Eubanks DC, Nelson MT,
JournalJ Biol Chem
PubMed ID12145283
'The nuclear factor of activated T-cells (NFAT), originally identified in T-cells, has since been shown to play a role in mediating Ca(2+)-dependent gene transcription in diverse cell types outside of the immune system. We have previously shown that nuclear accumulation of NFATc3 is induced in ileal smooth muscle by platelet-derived ... More
Application of a high-content multiparameter cytotoxicity assay to prioritize compounds based on toxicity potential in humans.
AuthorsAbraham VC, Towne DL, Waring JF, Warrior U, Burns DJ,
JournalJ Biomol Screen
PubMed ID18566484
'Prioritization of compounds based on human hepatotoxicity potential is currently a key unmet need in drug discovery, as it can become a major problem for several lead compounds in later stages of the drug discovery pipeline. The authors report the validation and implementation of a high-content multiparametric cytotoxicity assay based ... More
Entropic recoil separation of long DNA molecules.
AuthorsCabodi M, Turner SW, Craighead HG
JournalAnal Chem
PubMed ID12403567
'A novel technique that can rapidly separate long-strand polymers according to length is presented. The separation mechanism is mediated by a confinement-induced entropic force at the abrupt interface between regions of vastly different configuration entropy. To demonstrate this technique, DNA molecules were partially inserted into a dense array of nanopillars ... More
Intercalating fluorescence dye YOYO-1 prevents the folding transition in giant duplex DNA.
AuthorsYoshinaga N, Akitaya T, Yoshikawa K
JournalBiochem Biophys Res Commun
PubMed ID11500031
'Recently, it has become clear that with the addition of polyamines, giant DNA molecules of size greater than 10 kbp exhibit all-or-none switching between elongated coil and folded compact states. Here the effects of the intercalating fluorescent labeling dye, YOYO-1, and the minor-groove binding fluorescent labeling dye, DAPI, on the ... More
The yeast Sgs1 helicase is differentially required for genomic and ribosomal DNA replication.
AuthorsVersini G, Comet I, Wu M, Hoopes L, Schwob E, Pasero P
JournalEMBO J
PubMed ID12682026
'The members of the RecQ family of DNA helicases play conserved roles in the preservation of genome integrity. RecQ helicases are implicated in Bloom and Werner syndromes, which are associated with genomic instability and predisposition to cancers. The human BLM and WRN helicases are required for normal S phase progression. ... More
Optimisation of flow cytometric measurement of parasitaemia in plasmodium-infected mice.
AuthorsBarkan D, Ginsburg H, Golenser J
JournalInt J Parasitol
PubMed ID10779580
'Mouse malaria is often used as a model for drug testing. The results of drug trials are monitored by tedious (and consequently, sometimes inaccurate) microscopic counting of blood smears, or by flow cytometry. We suggest an improved, accurate and time-saving flow cytometric method for determination of parasitaemias in mice infected ... More
Differential activation of p70 and p85 S6 kinase isoforms during cardiac hypertrophy in the adult mammal.
AuthorsLaser M, Kasi VS, Hamawaki M, Cooper G, Kerr CM, Kuppuswamy D
JournalJ Biol Chem
PubMed ID9733756
'An adult feline right ventricular pressure overload (RVPO) model was used to examine the two S6 kinase (S6K) isoforms, p70(S6K) and p85(S6K), that are involved in translational and transcriptional activation. Biochemical and confocal microscopy analyses at the level of the cardiocyte revealed that p70(S6K) is present predominantly in the cytosol, ... More
Fluorometric determination of total mRNA with oligo(dT) immobilized on microtiter plates.
AuthorsMiura Y, Ichikawa Y, Ishikawa T, Ogura M, de Fries R, Shimada H, Mitsuhashi M
JournalClin Chem
PubMed ID8906073
'We have developed a rapid and nonradioactive method of quantifying cytosolic mRNA from crude cell lysates by using plastic plates to which oligonucleotides containing poly(dT) sequences were previously immobilized. Captured mRNA on the plate was mixed with Yoyo-1 fluorescent indicator dye, and the resulting Yoyo-1 fluorescence of the mRNA-Yoyo-1 complex ... More
Phase diagram, stability, and overcharging of lamellar cationic lipid-DNA self-assembled complexes.
AuthorsKoltover I, Salditt T, Safinya CR
JournalBiophys J
PubMed ID10423436
'Cationic lipid-DNA (CL-DNA) complexes comprise a promising new class of synthetic nonviral gene delivery systems. When positively charged, they attach to the anionic cell surface and transfer DNA into the cell cytoplasm. We report a comprehensive x-ray diffraction study of the lamellar CL-DNA self-assemblies as a function of lipid composition ... More
Diffusion of isolated DNA molecules: dependence on length and topology.
AuthorsRobertson RM, Laib S, Smith DE
JournalProc Natl Acad Sci U S A
PubMed ID16648255
'The conformation and dynamics of circular polymers is a subject of considerable theoretical and experimental interest. DNA is an important example because it occurs naturally in different topological states, including linear, relaxed circular, and supercoiled circular forms. A fundamental question is how the diffusion coefficients of isolated polymers scale with ... More
A rapid method for the preparation of single-cell suspensions from solid tissues.
AuthorsSingh NP
JournalCytometry
PubMed ID9515722
'A simple and rapid methodology for the preparation of single-cell suspension from animal tissues is described. It uses a device called a "tissue press" to mince the tissue into small pieces of desired sizes which are then dissociated into single cells using a pipette. The device is inexpensive and the ... More
Fluorometric nuclear run-on assay with oligonucleotide probe immobilized on plastic plates.
AuthorsOkamoto T, Mitsuhashi M, Kikkawa Y
JournalAnal Biochem
PubMed ID7985794
Stable dye-DNA intercalation complexes as reagents for high-sensitivity fluorescence detection.
AuthorsGlazer AN, Rye HS
JournalNature
PubMed ID1436062
'Fluorescent intercalation complexes of certain polycationic ligands with double-stranded DNA provide a new class of multichromophore labels for fluorescence assays.'
Imaging by fluorescence videomicroscopy of individual single stranded DNA molecules in solution.
AuthorsAuzanneau I, Barreau C, Salomé L
JournalC R Acad Sci III
PubMed ID8221228
'We demonstrate that the observation by fluorescence video-microscopy of individual single stranded DNA molecules is now possible owing to the use of an asymmetric cyanine (YOYO-1) belonging to a new class of fluorescent dyes complexing the DNA. The fluorescence of labelled DNA persists in denaturing conditions showing that the dye ... More
Real-time observation of a single DNA digestion by lambda exonuclease under a fluorescence microscope field.
'A fluorescence microscopy technique has been developed to visualize the behavior of individual DNA and protein molecules. Real-time direct observation of a single DNA molecule can be used to investigate the dynamics of DNA-protein interactions, such as the DNA digestion reaction by lambda exonuclease. In conventional methods it is impossible ... More
Semiautomated HLA-DQB1 typing by fluorescent dye photometry of amplified DNA on microtiter plates.
AuthorsBein G, Haase D, Schult J, Eiermann TH, Kirchner H
JournalHum Immunol
PubMed ID7910159
'We report a simple semiautomated HLA class II typing method that suits the demands of a 24-hour-duty transplantation service for preorgan retrieval donor typing. The procedure consists of sequence-specific amplification of HLA alleles by the polymerase chain reaction (nested PCR-SSP) followed by fluorescent dye photometry of the resulting PCR products ... More
Monitoring the amplification of CATCH, a 3SR based cooperatively coupled isothermal amplification system, by fluorimetric methods.
'Three different types of fluorescence detection methods were employed to monitor amplification of a previously established isothermal cooperatively coupled amplification system as it can serve as a tool for the investigation of fundamental issues in evolutionary optimization. By using 5''IRD-41 fluorescent labeled primers, the intercalating dye TOPRO-1 and a 5''fluorescin/3''DABCYL ... More
Clinical application of a novel DNA fluorescence intercalator, pyrylium dye, for the quantification of PCR products.
AuthorsIchikawa Y, Ishikawa T, Momiyama N, Chishima T, Hasegawa S, Yamaoka H, Yamamoto N, Okamoto T, Kawaguchi M, Shimada H
JournalScand J Clin Lab Invest
PubMed ID8981660
'Polymerase chain reaction (PCR) is a powerful tool for clinical diagnosis in the field of oncology, infection, and allergy. However, a simple and sensitive method for quantifying PCR products has not been established. We therefore used a novel fluorescence DNA intercalator, pyrylium iodide (P2) to quantify PCR products. A 838-bp ... More
The distribution of polycomb-group proteins during cell division and development in Drosophila embryos: impact on models for silencing.
AuthorsBuchenau P, Hodgson J, Strutt H, Arndt-Jovin DJ
JournalJ Cell Biol
PubMed ID9548724
'The subcellular three-dimensional distribution of three polycomb-group (PcG) proteins-polycomb, polyhomeotic and posterior sex combs-in fixed whole-mount Drosophila embryos was analyzed by multicolor confocal fluorescence microscopy. All three proteins are localized in complex patterns of 100 or more loci throughout most of the interphase nuclear volume. The rather narrow distribution of ... More
Novel expression of vascular endothelial growth factor receptor (VEGFR)-3 and VEGF-C on corneal dendritic cells.
AuthorsHamrah P, Chen L, Zhang Q, Dana MR
JournalAm J Pathol
PubMed ID12819011
'Vascular endothelial growth factor-3 (VEGFR-3) plays a critical role in embryonic cardiovascular development and is thought to be expressed exclusively on the lymphatic endothelium, high endothelial venules, and rarely on adult vascular endothelium. Recent evidence also suggests expression of VEGFR-3 on some tumor-associated macrophages. We have studied the expression of ... More
A plastid of probable green algal origin in Apicomplexan parasites.
'Protozoan parasites of the phylum Apicomplexa contain three genetic elements: the nuclear and mitochondrial genomes characteristic of virtually all eukaryotic cells and a 35-kilobase circular extrachromosomal DNA. In situ hybridization techniques were used to localize the 35-kilobase DNA of Toxoplasma gondii to a discrete organelle surrounded by four membranes. Phylogenetic ... More
Spectroscopic studies of the multiple binding modes of a trimethine-bridged cyanine dye with DNA.
AuthorsSovenyhazy KM, Bordelon JA, Petty JT
JournalNucleic Acids Res
PubMed ID12736305
'The interaction between DNA and a benzothiazole-quinoline cyanine dye with a trimethine bridge (TO-PRO-3) results in the formation of three noncovalent complexes. Unbound TO-PRO-3 has an absorption maximum (lambda(max)) of 632 nm, while the bound dyes (with calf thymus DNA) have electronic transitions with lambda(max) = 514 nm (complex I), ... More
KLP-18, a Klp2 kinesin, is required for assembly of acentrosomal meiotic spindles in Caenorhabditis elegans.
'The proper segregation of chromosomes during meiosis or mitosis requires the assembly of well organized spindles. In many organisms, meiotic spindles lack centrosomes. The formation of such acentrosomal spindles seems to involve first assembly or capture of microtubules (MTs) in a random pattern around the meiotic chromosomes and then parallel ... More
Real-time dynamics of single-DNA molecules undergoing adsorption and desorption at liquid-solid interfaces.
AuthorsKang SH, Shortreed MR, Yeung ES
JournalAnal Chem
PubMed ID11305636
'The conformational dynamics and adsorption/desorption behavior of individual lambda-DNA molecules at liquid-solid interfaces were monitored by imaging within the evanescent field layer using total internal reflection fluorescence microscopy. At a fused-silica surface, molecular conformation and adsorption behavior were found to depend on both pH and buffer composition. A histogram of ... More
Sequence of metabolic changes during X-ray-induced apoptosis.
AuthorsFerlini C, De Angelis C, Biselli R, Distefano M, Scambia G, Fattorossi A
JournalExp Cell Res
PubMed ID10047458
'A cell population undergoing apoptosis usually contains varying proportions of cells in the diverse stages of the process, from very early continuously through to secondary necrosis. This heterogeneity acts as a confounding factor in metabolic studies if a general population is investigated. Using fluorescent probes and multiparameter flow cytometry, we ... More
Fabrication of optically transparent carbon electrodes by the pyrolysis of photoresist films: approach to single-molecule spectroelectrochemistry.
AuthorsDonner S, Li HW, Yeung ES, Porter MD
JournalAnal Chem
PubMed ID16615798
'This paper describes the preparation, physical and chemical characterization, and performance of carbon-based optically transparent electrodes (C-OTEs) fabricated by the pyrolysis of thin films of photoresist. The electrodes are prepared by spin coating dilute solutions of the positive photoresist AZ 4330 onto quartz substrates. Pyrolysis of these samples at 1000 ... More
In situ screening assay for cell viability using a dimeric cyanine nucleic acid stain.
AuthorsBecker B, Clapper J, Harkins KR, Olson JA
JournalAnal Biochem
PubMed ID7527190
'A rapid and sensitive assay is described for the determination of cell viability of adherent and nonadherent cells that can be performed in situ in 96-well microtiter plates using fluorescence plate scanners. The assay, based on dye exclusion, utilizes a plasma membrane-impermeable, dimeric cyanine dye (YOYO-1). YOYO-1 fluoresces brightly only ... More
Inversions and gene order shuffling in Anopheles gambiae and A. funestus.
AuthorsSharakhov IV, Serazin AC, Grushko OG, Dana A, Lobo N, Hillenmeyer ME, Westerman R, Romero-Severson J, Costantini C, Sagnon N, Collins FH, Besansky NJ
JournalScience
PubMed ID12364797
'In tropical Africa, Anopheles funestus is one of the three most important malaria vectors. We physically mapped 157 A. funestus complementary DNAs (cDNAs) to the polytene chromosomes of this species. Sequences of the cDNAs were mapped in silico to the A. gambiae genome as part of a comparative genomic study ... More
Direct visualization of individual DNA molecules by fluorescence microscopy: characterization of the factors affecting signal/background and optimization of imaging conditions using YOYO.
AuthorsGurrieri S, Wells KS, Johnson ID, Bustamante C
JournalAnal Biochem
PubMed ID9193707
'A new series of high-affinity cyanine dyes was tested for the visualization of the dynamics of single DNA molecules through a fluorescence microscope. In particular, YOYO-1 (1,1''-(4,4,7,7-tetramethyl-4,7-diazaundecamethylene)- bis-4-[3-methyl-2,3-dihydro-(benzo-1,3-oxazole)- 2-methylidene]-quinolinium tetraiodide) forms a very stable, highly fluorescent complex with double-stranded DNA and dramatically improves the quality of the images. We have ... More
Direct visualization of abasic sites on a single DNA molecule using fluorescence microscopy.
AuthorsHirose T, Ohtani T, Muramatsu H, Tanaka A
JournalPhotochem Photobiol
PubMed ID12194206
'A new method was developed to allow direct visualization of damaged sites on individual DNA molecules. Fluorescence in situ hybridization on extended DNA molecules was modified to detect a single abasic site. Abasic sites were specifically labeled with a biotinylated aldehyde-reactive probe and fluorochrome-conjugated streptavidin. The light emitted by a ... More
Packaging of single DNA molecules by the yeast mitochondrial protein Abf2p.
AuthorsBrewer LR, Friddle R, Noy A, Baldwin E, Martin SS, Corzett M, Balhorn R, Baskin RJ
JournalBiophys J
PubMed ID14507714
'Mitochondrial and nuclear DNA are packaged by proteins in a very different manner. Although protein-DNA complexes called "nucleoids" have been identified as the genetic units of mitochondrial inheritance in yeast and man, little is known about their physical structure. The yeast mitochondrial protein Abf2p was shown to be sufficient to ... More
The use of fluorescence enhancement to improve the microscopic diagnosis of falciparum malaria.
AuthorsGuy R, Liu P, Pennefather P, Crandall I,
JournalMalar J
PubMed ID17617912
'BACKGROUND: Giemsa staining of thick blood smears remains the "gold standard" for detecting malaria. However, this method is not very good for diagnosing low-level infections. A method for the simultaneous staining of Plasmodium-parasitized culture and blood smears for both bright field and fluorescence was developed and its ability to improve ... More
Scanning force microscopy of DNA molecules elongated by convective fluid flow in an evaporating droplet.
AuthorsWang W, Lin J, Schwartz DC
JournalBiophys J
PubMed ID9649413
'Scanning force microscopy (SFM) was used to image intact, nearly fully elongated lambda bacteriophage DNA molecules, fixed onto freshly cleaved mica surfaces. Molecular elongation and fixation were accomplished using a newly characterized fixation technique, termed "fluid fixation." Here convective fluid flows generated within an evaporating droplet of DNA solution efficiently ... More
Fluorometric assay using dimeric dyes for double- and single-stranded DNA and RNA with picogram sensitivity.
AuthorsRye HS, Dabora JM, Quesada MA, Mathies RA, Glazer AN
JournalAnal Biochem
PubMed ID7679561
'Thiazole orange homodimer (TOTO; 1,1''-(4,4,7,7-tetramethyl-4,7-diazaundecamethylene)-bis-4-[3-methy l-2,3- dihydro-(benzo-1,3-thiazole)-2-methylidene]-quinolinium tetraiodide) and oxazole yellow homodimer (YOYO; an analogue of TOTO with a benzo-1,3-oxazole in place of the benzo-1,3-thiazole) bind with very high affinity to nucleic acids with more than a 1000-fold fluorescence enhancement upon binding. A linear dependence of fluorescence intensity on DNA ... More
Photobleaching of asymmetric cyanines used for fluorescence imaging of single DNA molecules.
AuthorsKanony C, Akerman B, Tuite E
JournalJ Am Chem Soc
PubMed ID11506554
'The photobleaching of the cyanine dyes YO and YOYO has been investigated for both free and DNA-bound dyes, using absorption and fluorescence spectroscopy coupled with fluorescence microscopy. For the free dyes, the nature of the reactive species involved in the photodegradation process is different for the monomer and the dimer, ... More
Direct reverse transcription-PCR on oligo(dT)-immobilized polypropylene microplates after capturing total mRNA from crude cell lysates.
AuthorsHamaguchi Y, Aso Y, Shimada H, Mitsuhashi M
JournalClin Chem
PubMed ID9799751
'To simplify gene expression analysis, oligo(dT)-immobilized polypropylene microplates were used serially to capture mRNA, synthesize cDNA, and amplify specific genes. The amounts of immobilized oligonucleotide, hybridized mRNA, and synthesized cDNA were quantified fluorometrically using either Yoyo-1 or AttoPhos. The immobilized oligonucleotides captured approximately 40-55% of mRNA directly from crude cell ... More
Nuclear organization studied with the help of a hypotonic shift: its use permits hydrophilic molecules to enter into living cells.
AuthorsKoberna K, Stanek D, Malínský J, Eltsov M, Pliss A, Ctrnáctá V, Cermanová S, Raska I
JournalChromosoma
PubMed ID10525969
'A new procedure for introduction of hydrophilic molecules into living cells based on efficient uptake of these molecules into the cells during hypotonic treatment is presented and its use is demonstrated by a variety of applications. Experiments with cultured vertebrate and Drosophila cells and various animal tissues demonstrated that the ... More
Structural analyses of purified human immunodeficiency virus type 1 intracellular reverse transcription complexes.
AuthorsNermut MV, Fassati A
JournalJ Virol
PubMed ID12857888
'Retroviruses copy their RNA genome into a DNA molecule, but little is known of the structure of the complex mediating reverse transcription in vivo. We used confocal and electron microscopy to study the structure of human immunodeficiency virus type 1 (HIV-1) intracellular reverse transcription complexes (RTCs). Cytoplasmic extracts were prepared ... More
One-end immobilization of individual DNA molecules on a functional hydrophobic glass surface.
AuthorsMatsuura S, Kurita H, Nakano M, Komatsu J, Takashima K, Katsura S, Mizuno A
JournalJ Biomol Struct Dyn
PubMed ID12437381
'We demonstrate an effective method for DNA immobilization on a hydrophobic glass surface. The new DNA immobilizing technique is extremely simple compared with conventional techniques that require heterobifunctional crosslinking reagent between DNA and substrate surface that are both modified chemically. In the first process, a coverslip was treated with dichlorodimethylsilane ... More
Cortical plaque-like structures identify ribosome-containing domains in the Mauthner cell axon.
AuthorsKoenig E, Martin R
JournalJ Neurosci
PubMed ID8778291
'The hypothesis that ribosomes are present, but may have a restricted distribution, in the Mauthner (M) axon was evaluated in isolated M-cell axoplasm after (1) staining with YOYO-1 and (2) inspection by electron spectroscopic imaging (ESI) of ribosomal RNA (rRNA) phosphorus (P). Discrete periaxoplasmic plaques, identified by their ribonuclease-sensitive fluorescence, ... More
Visual quantification of DNA double-strand breaks in bacteria.
AuthorsSingh NP, Stephens RE, Singh H, Lai H
JournalMutat Res
PubMed ID10526201
'In this paper, we describe a method for the visualization of double-strand breaks in a single electrostretched Escherichia coli DNA molecule. We also provide evidence that electrostretched or migrated DNA under neutral microgel electrophoresis conditions is made up of individual chromosomes. Using the neutral microgel electrophoresis technique, DNA migration (stretching) ... More
Rapid DNA fiber technique for size measurements of linear and circular DNA probes.
AuthorsHenegariu O, Grober L, Haskins W, Bowers PN, State MW, Ohmido N, Bray-Ward P, Ward DC
JournalBiotechniques
PubMed ID11515354
Comet assay coupled to repair enzymes for the detection of oxidative damage to DNA induced by low doses of gamma-radiation: use of YOYO-1, low-background slides, and optimized electrophoresis conditions.
Optical mapping: a novel, single-molecule approach to genomic analysis.
AuthorsSamad A, Huff EF, Cai W, Schwartz DC
JournalGenome Res
PubMed ID8717049
Use of the fluorescent dye YOYO-1 to quantify oligonucleotides immobilized on plastic plates.
AuthorsOgura M, Keller C, Koo K, Mitsuhashi M
JournalBiotechniques
PubMed ID8074866
Fluorometric method for the measurement of nuclease activity on plastic plates.
AuthorsOgura M, Mitsuhashi M
JournalBiotechniques
PubMed ID7727122
Screening method for a large quantity of polymerase chain reaction products by measuring YOYO-1 fluorescence on 96-well polypropylene plates.
AuthorsOgura M, Mitsuhashi M
JournalAnal Biochem
PubMed ID8074307
Staining of DNA in live and fixed cells.
AuthorsCrissman HA, Hirons GT
JournalMethods Cell Biol
PubMed ID7532262
Use of nucleic acid dyes SYTO-13, TOTO-1, and YOYO-1 in the study of Escherichia coli and marine prokaryotic populations by flow cytometry.
AuthorsGuindulain T, Comas J, Vives-Rego J
JournalAppl Environ Microbiol
PubMed ID9361447
Three nucleic acid dyes (SYTO-13, TOTO-1, and YOYO-1) were tested on cultures of Escherichia coli and marine prokaryote populations. These dyes stain the RNA and DNA in E. coli but only respond to DNA in marine populations, according to the histograms obtained after DNase and RNase treatments. ... More
Single-molecule denaturation mapping of DNA in nanofluidic channels.
AuthorsReisner W, Larsen NB, Silahtaroglu A, Kristensen A, Tommerup N, Tegenfeldt JO, Flyvbjerg H,
JournalProc Natl Acad Sci U S A
PubMed ID20616076
Here we explore the potential power of denaturation mapping as a single-molecule technique. By partially denaturing YOYO-1-labeled DNA in nanofluidic channels with a combination of formamide and local heating, we obtain a sequence-dependent ... More
Methods and applications of three-color confocal imaging.
AuthorsPaddock SW, Hazen EJ, De Vries PJ
JournalBiotechniques
PubMed ID8994659
A simple method for constructing two- and three-color merged images from grayscale confocal fluorescence images using Adobe Photoshop is outlined. Various computer methods for manipulating and displaying the images are discussed in light of several recent biomedical applications of multi-label confocal microscopy. ... More
Encoded reaction cassette for the highly sensitive detection of the making and breaking of chemical bonds.
AuthorsFenniri H, Janda KD, Lerner RA
JournalProc Natl Acad Sci U S A
PubMed ID7892261
A reaction cassette has been designed for the highly sensitive detection of the making and breaking of chemical bonds. The system is envisioned as a companion device to be used in the search for antibody and other novel catalysts. The cassette also may have important clinical applications in the design ... More