Protein microarrays antibody

Because protein microarrays (antibody microarrays) have been successfully used for a variety of applications, such as antibody response profiling, identification and detection of bacterial and protein analytes, as well as disease proteomics with a clear focus toward oncoproteomics, it can be assumed that they will also prove to be a useful tool in studies on allergens (Wingren and Borrebaeck, 2007). 

Keywords Microarray DNA microarray Carbohydrate microarray Protein microarray Antibody microarray G protein-coupled receptor microarray Cellular microarray Optical biosensor Resonant waveguide grating biosensor Surface plasmon resonance Dynamic mass redistribution... 

Figure 7.7. Protein microarray on polyvinylidene difluoride filters (PVDF). Proteins are gridded on filter and probed with an antibody to a gene of interest. Protein-ligand interactions could be detected using a labeled ligand as a probe.

The potential of photolithography for the construction of protein microarrays has also been demonstrated (Mooney et al., 1996). For these experiments, antibodies were assembled in precise two-dimensional patterns on silicon wafers. This was accomplished by first forming a self-assembled monolayer of n-octadecyltrimethoxysilane (OTMS) on a silicon-dioxide... 

Lueking, A., Horn, M., Eickhoff, H., Bussow, K., Lehrach, H., and Walter, G. (1999). Protein microarrays for gene expression and antibody screening. Anal. Biochem. 270, 103-111. 

Peluso P., Wilson D.S., Do D., Tran H., Venkatasubbaiah M., Quincy D., Heidecker B., Poindexter K., Tolani N., Phelan M., Witte K., Jung L.S., Wagner P., Nock S., Optimizing antibody immobilization strategies for the construction of protein microarrays, Anal Biochem. 2003 312 113-124. 

Currently, protein microarrays can be divided into various types depending on the strategies to be chosen. For example, according to the array structure and shape, protein microarrays include 3D-surface structure [30, 31], nanowell [32], and plain chips [33-36], Meanwhile, considering the field of application, protein microarrays can be classified into five categories antibody array, antigen or reserve array, functional array, capture array, and solute array. Table 11.6 shows the differences among them. 

The most common use of protein microarrays is in immunoassays. In particular, antibody-based immunoassays are the main stream of diagnostic assays due to their specificity. The assay usually runs in a multiplexed mode where the antibodies or other capture agents are immobilized and then exposed to a biological sample. There are four immunoassay formats direct binding, sandwich (ELISA), competitive, and displacement. Direct-binding and sandwich assays are the most common. There are some reports on the use of competitive assays and displacement assays, which are usually associated with high surface area/volume systems [72-76],... 

Wang, X. Zhao, M. Nolte, D. D., Area scaling of interferometric and fluoresecnt detection of protein on antibody microarrays, Biosen. Bioelectron. 2008... 

Haab, B.B., Dunham, M.J., and Brown, P.O., Protein microarrays for highly parallel detection and quantitation of specific proteins and antibodies in complex solutions. Genome Biol. 2, 0004.1-0004.13, 2001. 

Probes can be antibodies, other binding proteins constructed from protein fusions, or even oligonucleotide aptamers. While completion of the Human Genome Project has enabled access to content for nuclide acid arrays, the content for protein arrays is largely based upon available antibody libraries. Thus, the commercialization of protein microarrays remains largely dependent upon both commercial and institutional providers of protein content. These providers must also permit access to the data-based protein annotations. These are necessary in order for the protein array to be useful as a bioinformatics tool. 

Novagen (ProteoPlex ), S S (FAST Quant), BioSource (Cartesian Array ), and BD Biosciences (BD Clontech Ab Microarrays) have introduced or will soon introduce protein microarray slide formatted products in which antibodies are directly immobilized. Beckman Coulter s protein array products for performing micro-ELlSAS in standard 96-well plate formats are based upon the self-assembly (by hybridization) of oligonucleotide-antibody conjugates to complementary oligonucleotides arrayed in individual wells. HTG s protein array technology was described previously. 

The manufacture and processing of the protein microarray should be conducted in such a manner that the arrayed proteins remain in their native and active state. For most proteins, this usually means the hydrated state in order to avoid surface denaturation. For antibody arrays which are perhaps more forgiving than other proteins, it has been our experience that while these could be stored cold and dry, it is most important to rehydrate them prior to use. This process is in sharp contrast to the preparation of nucleic acid arrays in which strand melting or denaturahon is necessary to achieve optimal binding to the solid support. While the hybridization process is well understood and can be controlled under thermodynamic principles, the folding and renaturation of proteins on planar (microarray) surfaces is under study. 

Such thermoplastics have also been used as DNA microarray substrates (Matson et al., 1995 Shchepinov, 1997 Beier and Hoheisel, 1999) and in the construction of protein microarrays in microwells (Matson et al., 2001 Moody, 2001). Pierce (see searchlight perbio.com) introduced the Search-Light series of microarray-based ELISA assays immobilizing capture antibodies in a low density array format into polystyrene microwells. 

Substrates for the creation of protein microarrays were initially selected from those used for DNA arrays, for example, PLL glass slides. At first, these substrates proved to be sufficient for antibody microarray studies. However, not all proteins will behave well or similarly on a particular substrate material. New solid phases applicable for protein microarrays need to be found. 

Angenendt, R, Glokler, J., Sobek, J., Lehrach, H., and Cahill, D.J., Next generation of protein microarray support materials Chromatography A 1009 97-104, 2003. evaluation for protein and antibody microarray applications. 

Huang, R.P, Detection of multiple proteins in an antibody-based protein microarray system, /. Immunol. Methods, 255, 1-13, 2001. 

Angenendt et al. (2003) evaluated several slide surface chemistries for use as protein and antibody microarrays. They reasoned that because proteins vary greatly in their surface charge and relative hydrophobicity, a careful selection of surface chemistry may be important for obtaining optimal performance for a particular protein. Thus, several commercially available slide surface chemistries were evaluated for performance in model arrays... 

The protein microarray represents an emerging technology. While we have described its potential utility, several key problems remain to be overcome before this tool is fully adopted by the research and biopharmaceutical commxmities. The most likely first embodiment will be an antibody "protein-detecting" microarray. This is understandable based upon the availability and suitability of antibody libraries originally developed for ELISA. We have discussed many demonstrahons of antibody arrays in this chapter but commercial introductions (Pierce, Beckman Coulter) have been limited. 

A. Lucking, M. Horn, H. Eickhoff, K. Bussow, H. Lehrach, and G. Walter. 1999. Protein Microarrays for Gene Expression and Antibody Screening. Analytical Biochemistry 270 103-111. 

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