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The coupling of quantum dots (QDs) to antibodies heralds a new era of versatility in multicolor flow cytometry. This chapter introduces the properties of QDs (as relevant to flow cytometric applications), the advantages derived from these properties, and the procedure for conjugating a ...

This chapter describes the methodology and a detailed protocol for the targeting and optical imaging of experimental brain tumors in rats using quantum dots (QDs). QDs are optical semiconductor nanocrystals that exhibit stable, bright fluorescence over narrow, size-tunable emis ...

The development of new fluorophores has experienced a tremendous advance over the last two decades. The unique photophysical properties of quantum dots (QDs), such as their large Stokes shifts and exceptional brightness, make them attractive probes in flow cytometry applications. In ...

We have developed several conjugation strategies based on noncovalent self-assembly for the attachment of proteins and other biomolecules to water-soluble luminescent colloidal semiconductor nanocrystals (quantum dots ). The resulting QD-protein conjugates were empl ...

Reverse-phase protein microarrays (RPPMAs) enable heterogeneous mixtures of proteins from cellular extracts to be directly spotted onto a substrate (such as a protein biochip) in minute volumes (nanoliter-to-picoliter volumes). The protein spots can then be probed with primary a ...

Highly multiplexed genomics assays are challenged by the need for a sufficient signal-to-noise ratio for each marker scored on a microarray-detection platform. Typically, as the number of markers scored (or target complexity) increases, either more assay-target material must be app ...

Fluorescence is highly sensitive to environment, and the distance separating fluorophores and quencher molecules can provide the basis for effective homogeneous nucleic acid hybridization assays. Molecular interactions leading to fluorescence quenching include coll ...

Rapid and reliable detection of mutations at the genetic level is an integral part of modern molecular diagnostics. These mutations can range from dominant single nucleotide polymorphisms within specific loci to codominant heterozygotic insertions and they present considera ...

Conventional methods to detect Salmonella spp. in foodstuffs may take up to 1 wk. Methods for pathogen detection are required. Real-time detection of Salmonella spp. will broaden our ability to screen large number of samples in a short time. This chapter describes a step-by-step procedure using ...

The 26S proteasome is the executing protease of the ubiquitin-dependent degradation system. It consists of one or two 19S regulatory sub-complexes and one 20S proteolytic sub-complex (1). The 20S proteasome is a barrel-shaped cylinder which consists or four stacked rings (2). Each of the two o ...

Sensitive TaqMan� real-time polymerase chain reaction (PCR)-based methods have been developed recently for the detection and quantitation of feline leukemia virus (FeLV) proviral DNA in infected cats. In this chapter, we outline the design and implementation of a TaqMan� real-time PCR ...

Due to the fact that mutations and up- or downregulation of genes can lead to the development of cancer, quantitative comparison of relative gene expression in healthy and cancerous tissue can gain valuable insights into tumorigenesis. While the semi-quantitative DNA microarrays are be ...

Real-time PCR is presently the gold standard of gene expression quantification. Configuration of real-time PCR instruments with 384-well reaction blocks, enables the instrument to be used essentially as a low-density array. While PCR will never rival the throughput of microchip arra ...

A number of probe systems exist for the real-time detection of PCR products. Scorpions are a unique method wherein primer and probe are combined in a single oligonucleotide. During the PCR, the probe element becomes linked directly to its complementary target site with beneficial consequen ...

Real-time polymerase chain reaction (PCR) has become a standard tool in both quantitative gene expression and genetic variation analysis. Data collection is performed throughout the PCR process, thus combining amplification and detection into a single step. This can be achieved by co ...

This chapter describes two different fast protocols for optimizing EasyBeacons™ for the challenging detection of the methylation status of a single CpG duplet in bisulfitetreated DNA. EasyBeacons™ can be used in multiplex detections even if they do not have the same affinity for their res ...

HyBeacon probes are single-stranded oligonucleotides with one or more internal base(s) labeled with a fluorescent dye. When a probe forms a duplex with its target sequence, the level of fluorescence emission increases considerably. HyBeacons have been developed as new tools for rapid se ...

Methods are described for preparation and use of quenched autoligation (QUAL) probes. These modified oligonucleotide fluorescent probes can be used to detect DNA and RNA in solution, on solid surfaces, and in fixed and living bacterial and human cells. They are quenched probes, and thus provi ...

Forced intercalation probes (FIT-probes) are peptide nucleic acid-based probes in which the thiazole orange dye replaces a canonical nucleobase. FIT-probes are used in homogenous DNA detection. The analysis is based on sequence-specific binding of the FIT-probe with DNA. Binding of t ...

Fluorescent nucleic acid detection in polymerase chain reaction (PCR) generally uses oligonucleotide probes labeled with covalently attached dyes. However, unlabeled oligonucleotides in the presence of saturating DNA dyes can also serve as hybridization probes. The DNA dye, L ...