Abstract
The generation of recombinant antibodies by phage display in high-throughput demands fast downstream technologies and streamlined processes for the identification and initial characterisation of individual binders. Next to standard immunological methods such as enzyme-linked immunosorbent assays (ELISA) and Western-blot, protein microarrays offer a wide range of possibilities in the evaluation process of monoclonal binders. Here, we describe the application of a special protein microarray method – the multiple spotting technique (MIST) – for the simultaneous evaluation of hundreds of phage display derived soluble monoclonal antibody fragments on protein microarrays. The standard operating procedures provided include the expression of soluble antibody fragments in microtitre plates, the spotting protocols and data evaluation schemes. Additionally, we show the comparability of this protein microarray application to conventional ELISA on a recent target antigen in our semi-automated selection pipeline. Applying MIST allows to reduce time, material and waste, and extends automation beyond the selection process applying conventional microarray machinery.
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Acknowledgements
This work was supported by the German Federal Ministry for Education and Research (BMBF) through the National Genome Research Network (NGFN-II) project “Antibody Factory” (Grant No. 01GR0427) and the Max Planck Society. ZK acknowledges additional support from EU-FP6 CA “Proteome Binders” (RICA 026008).
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Konthur, Z., Wilde, J. (2010). Evaluation of Recombinant Antibodies on Protein Microarrays Applying the Multiple Spotting Technique. In: Kontermann, R., Dübel, S. (eds) Antibody Engineering. Springer Protocols Handbooks. Springer, Berlin, Heidelberg. https://doi.org/10.1007/978-3-642-01147-4_34
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DOI: https://doi.org/10.1007/978-3-642-01147-4_34
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