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Re: protien chips
Protein microarray technology offers direct detection and quantification of protein expression, the endpoint of both molecular and cellular function in health and disease. Protein microarrays have been used for elucidation of protein function and signaling, protein-protein interactions, detection of bacteria and toxins, drug discovery, and identification of protein biomarkers. Protein-binding microarrays, typically containing spotted antibodies, has been used to profile protein expression levels in complex samples such as crude cell lysates and serum. In these cases, the limits of detection reached the low ng/mL level, given sufficiently specific antibodies. Unfortunately, this technology is less sensitive than comparable sandwich immunoassay-based assays, but sufficient to detect at moderately expressed proteins.
Protein microarrays exist in two general formats, the forward-phase array (FPAs) and the reverse-phase protein microarray (RPMAs). FPAs are constructed by printing many bait molecules (such as antibodies) onto an array surface, allowing for the profiling of complex samples; such arrays are usually probed with cell lysates or patient serum samples. RPAs, however, are constructed by printing multiple test samples and can be probed using a single detector, such as an antibody. While the reverse-phase format was originally used to identify biomarkers in laser capture microdissection (LCM) samples, RPAs can be constructed using a wide variety of test samples including serum, bodily fluids (cerebrospinal fluids, synovial fluid, etc.), cell lysate, and protein fractions.
Protein immobilization on a nitrocellulose substratum occurs via multiple, poorly understood interactions consisting of van der Waals, electrostatic, and hydrophobic interactions. The common element for each type of microarray is the immobilization of bait molecules on a substratum, avoiding evaporation during the printing process, either as a homogeneous or heterogeneous spot. Protein detection strategies foresee, specific high affinity antibodies and ligands as probe molecules and high specificity and sensitivity system for detect the complex by means chemistries amplification techniques with chromogenic (DAB, TMB, Luminol, etc.) and fluorometric (Cy3, Cy5, etc.) detection of proteins. Enzyme chromogenic oxidation conducts to the formation of precipitate product that is a permanent signals with femtomolar sensitivity, low background, and easily visualized. The level of sensitivity is necessary for applications related to biological response monitoring small volume samples such as laser-capture microdissected samples. The antibody affinity also determines the assay linearity range. Linearity can only be achieved when the concentrations of the analyte and antibody are matched to the affinity constant. Unfortunately, multiplexed formats containing multiple antibodies with varying affinities will not be able to achieve linearity for all analytes in each spot. If you desire to know an way for to realize a home made protein microarray system on microscope slide glass, I is available for it, while for more to know see : Methods in Molecular Biology - Vol. 671 - Biological Microarrays Methods and Protocols.
|chips , protien|
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