Summary
This chapter describes the development and use of bead-based miniaturized multiplexed sandwich immunoassays for focused protein profiling. Bead-based protein arrays or suspension microarrays allow simultaneous analysis of a variety of parameters within a single experiment. In suspension microarrays capture antibodies are coupled onto color-coded microspheres.
The applications of suspension microarrays are described, which allow to analyze proteins present in different types of body fluids, such as serum or plasma, cerebrospinal, pleural and synovial fluids, as well as cell culture supernatants. The chapter is divided into the generation of suspension microarrays, sample preparation, processing of suspension microarrays, validation of analytical performance, and finally pattern generation using bioinformatics tools.
This is a preview of subscription content, log in via an institution.
Buying options
Tax calculation will be finalised at checkout
Purchases are for personal use only
Learn about institutional subscriptionsSpringer Nature is developing a new tool to find and evaluate Protocols. Learn more
References
Morgan, E., Varro, R., Sepulveda, H., Ember, J.A., Apgar, J., Wilson, J., Lowe, L., Chen, R., Shivraj, L., Agadir, A., Campos, R., Ernst, D., Gaur, A. (2004) Cytometric bead array: a multiplexed assay platform with applications in various areas of biology. Clin Immunol, 110, 252–66
Dasso, J., Lee, J., Bach, H., Mage, R.G. (2002) A comparison of ELISA and flow microsphere-based assays for quantification of immunoglobulins. J Immunol Methods, 263, 23–33
Carson, R.T., Vignali, D.A. (1999) Simultaneous quantitation of 15 cytokines using a multiplexed flow cytometric assay. J Immunol Methods, 227, 41–52
Dunbar, S.A., Vander Zee C.A., Oliver, K.G., Karem, K.L., Jacobson, J.W. (2003). Quantitative, multiplexed detection of bacterial pathogens: DNA and protein applications of the Luminex LabMAP system. J Microbiol Methods, 53, 245–52
Joos, T.O., Stoll, D., Templin, M.F. (2002) Miniaturised multiplexed immuno- assays. Curr Opin Chem Biol, 6, 76–80
Prabhakar, U., Eirikis, E., Davis, H.M. (2002) Simultaneous quantification of proinflammatory cytokines in human plasma using the LabMAP assay. J Immunol Methods, 260, 207–18
Lu, J., Getz, G., Miska, E.A., Alvarez-Saavedra, E., Lamb, J., Peck, D., Sweet-Cordero, A., Ebert, B.L., Mak, R.H., Ferrando, A.A., Downing, J.R., Jacks, T., Horvitz, H.R., Golub, T.R. (2005) MicroRNA expression profiles classify human cancers. Nature, 435, 834–8
de Jager, W., Prakken, B.J., Bijlsma, J.W., Kuis, W., Rijkers, G.T. (2005) Improved multiplex immunoassay performance in human plasma and synovial fluid following removal of interfering heterophilic antibodies. J Immunol Methods, 300, 124–35
Natelson, B.H., Weaver, S.A., Tseng, C.L., Ottenweller, J.E. (2005) Spinal fluid abnormalities in patients with chronic fatigue syndrome. Clin Diagn Lab Immunol, 12, 52–5
Findlay, J.W., Smith, W.C., Lee, J.W., Nordblom, G.D., Das, I., DeSilva, B.S., Khan, M.N., Bowsher, R.R. (2000) Validation of immunoassays for bioanalysis: a pharmaceutical industry perspective. J Pharmaceutical Biomed Anal, 21, 1249–73
Sanchez-Carbayo, M. (2006) Antibody arrays: technical considerations and clinical applications in cancer. Clin Chem, 52, 1651–9
Kingsmore, S.F. (2006) Multiplexed protein measurement: technologies and applications of protein and antibody arrays. Nat Rev Drug Discov, 5, 310–20
Weinstein, J.N., Myers, T.G., O’Connor, P.M., et al. (1997) An information-intensive approach to the molecular pharmacology of cancer. Science, 275, 343–9
Saeed, A.I., Sharov, V., White, J., Li, J., Liang, W., Bhagabati, N., Braisted, J., Klapa, M., Currier, T., Thiagarajan, M., Sturn, A., Snuffin, M., Rezantsev, A., Popov, D., Ryltsov, A., Kostukovich, E., Borisovsky, I., Liu, Z., Vinsavich, A., Trush, V., Quackenbush, J. (2003). TM4: a free, open-source system for microarray data management and analysis. Biotechniques, 34(2), 374–8.
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2008 Humana Press, a part of Springer Science+Business Media, LLC
About this protocol
Cite this protocol
Hsu, HY., Wittemann, S., Joos, T.O. (2008). Miniaturized Parallelized Sandwich Immunoassays. In: Vlahou, A. (eds) Clinical Proteomics. Methods in Molecular Biology™, vol 428. Humana Press. https://doi.org/10.1007/978-1-59745-117-8_14
Download citation
DOI: https://doi.org/10.1007/978-1-59745-117-8_14
Publisher Name: Humana Press
Print ISBN: 978-1-58829-837-9
Online ISBN: 978-1-59745-117-8
eBook Packages: Springer Protocols