Functional protein microarrays also known as target protein arrays are constructed by immobilising large numbers of purified proteins and are used to identify protein—protein, protein—DNA, protein— RNA , protein— phospholipid , and protein—small-molecule interactions, to assay enzymatic activity and to detect antibodies and demonstrate their specificity.
They differ from analytical arrays in that functional protein arrays are composed of arrays containing full-length functional proteins or protein domains. These protein chips are used to study the biochemical activities of the entire proteome in a single experiment. The key element in any functional protein microarray-based assay is the arrayed proteins must retain their native structure, such that meaningful functional interactions can take place on the array surface.
The advantages of controlling the precise mode of surface attachment through use of an appropriate affinity tag are that the immobilised proteins will have a homogeneous orientation resulting in a higher specific activity and higher signal-to-noise ratio in assays, with less interference from non-specific interactions. Protein array detection methods must give a high signal and a low background.
Protein microarray - Wikipedia
The most common and widely used method for detection is fluorescence labeling which is highly sensitive, safe and compatible with readily available microarray laser scanners. Other labels can be used, such as affinity, photochemical or radioisotope tags. These labels are attached to the probe itself and can interfere with the probe-target protein reaction.
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Therefore, a number of label free detection methods are available, such as surface plasmon resonance SPR , carbon nanotubes, carbon nanowire sensors where detection occurs via changes in conductance and microelectromechanical system MEMS cantilevers. All these label free detection methods are relatively new and are not yet suitable for high-throughput protein interaction detection; however, they do offer much promise for the future. Protein quantitation on nitrocellulose coated glass slides can use near-IR fluorescent detection. This limits interferences due to auto-fluorescence of the nitrocellulose at the UV wavelengths used for standard fluorescent detection probes.
Protein Arrays, Biochips and Proteomics: The Next Phase of Genomic Discovery (No Series)
There are five major areas where protein arrays are being applied: diagnostics, proteomics, protein functional analysis, antibody characterization, and treatment development. Diagnostics involves the detection of antigens and antibodies in blood samples; the profiling of sera to discover new disease biomarkers ; the monitoring of disease states and responses to therapy in personalized medicine; the monitoring of environment and food.
Digital bioassay is an example of using protein microarray for diagnostic purposes. A cost-effective fabrication platform using OSTE polymers for such microwell arrays has been recently demonstrated and the bio-assay model system has been successfully characterised. Proteomics pertains to protein expression profiling i.
Protein functional analysis is the identification of protein—protein interactions e. Antibody characterization is characterizing cross-reactivity , specificity and mapping epitopes. Treatment development involves the development of antigen-specific therapies for autoimmunity , cancer and allergies; the identification of small molecule targets that could potentially be used as new drugs. Despite the considerable investments made by several companies, proteins chips have yet to flood the market.
Manufacturers have found that proteins are actually quite difficult to handle. Production of reliable, consistent, high-throughput proteins that are correctly folded and functional is fraught with difficulties as they often result in low-yield of proteins due to decreased solubility and formation of inclusion bodies. There are a number of approaches to this problem which differ fundamentally according to whether the proteins are immobilised through non-specific, poorly defined interactions, or through a specific set of known interactions. The former approach is attractive in its simplicity and is compatible with purified proteins derived from native or recombinant sources   but suffers from a number of risks.
Most notable amongst these relate to the uncontrolled nature of the interactions between each protein and the surface; at best, this might give rise to a heterogeneous population of proteins in which active sites are sometimes occluded by the surface; at worst, it might destroy activity altogether due to partial or complete surface-mediated unfolding of the immobilised protein. Challenges include: 1 finding a surface and a method of attachment that allows the proteins to maintain their secondary or tertiary structure and thus their biological activity and their interactions with other molecules, 2 producing an array with a long shelf life so that the proteins on the chip do not denature over a short time, 3 identifying and isolating antibodies or other capture molecules against every protein in the human genome, 4 quantifying the levels of bound protein while assuring sensitivity and avoiding background noise, 5 extracting the detected protein from the chip in order to further analyze it, 6 reducing non-specific binding by the capture agents, 7 the capacity of the chip must be sufficient to allow as complete a representation of the proteome to be visualized as possible; abundant proteins overwhelm the detection of less abundant proteins such as signaling molecules and receptors, which are generally of more therapeutic interest.
From Wikipedia, the free encyclopedia. Bibcode : Natur. Protein Microarrays. Nature Biotechnology. Ageing Dev. Chemistry of Materials.
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However, as biochip systems comprise probes formatted in micro scales on glass surfaces, they require automated robotic tools for handling and scanning of micro-scale samples. In addition, bioinformatics tools are also needed for data analysis. Need for such specific tools and techniques for implementation of this technology is expected to make biochips a highly sophisticated and considerably expensive technology, along with the expansion of the product range for the market.
Simultaneous and specific profiling of biological markers is a major application of biochips. In addition, they are also critical components in studies investigating multiple parameters simultaneously, as they hold chemical sites providing specific and sensitive analysis.
Furthermore, these integrated systems involve exhaustive physical and chemical analysis for studies related to optimal selection, activation, and stabilization for the development of new platforms of clinical systems. DNA chips accounted for the largest share of revenue, as a result of high spending on research pertaining to genomics and next-generation sequencing along with rising demand for detection and diagnosis of genetic disorders.
Progress in the development of protein microarrays coupled with emerging classes of tissue- and cell-based biochips for various applications are factors anticipated to drive demand for biochips.
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Also, the competitive dynamics prevalent are greatly influenced by next-generation sequencing applications. Applications to enhance information support and lower the cost of miniaturized bioanalysis are anticipated to enhance biochip demand over the forecast period. The protein and DNA biochips and LOACs have widespread applications for gene expression, single nucleotide polymorphism genotyping, expression profiling, in vitro diagnosis, high throughput screening, point-of-care, drug discovery, tissue arrays, and agricultural biotechnology.
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