Abstract
This simple protocol describes how to detect antigens from agar-grown bacterial colonies transferred to nitrocellulose using specific antibodies. The protocol is well suitable for detection of bacterial proteins exposed on the cell surface or secreted to the extracellular space and it can be modified also for detection of intracellular proteins. The assay can distinguish bacterial clones with different expression rates (high, medium and low) from colonies that do not express target protein. We used this assay for screening of Mat fimbriae-producing Escherichia coli mutants obtained by mini-Tn5 transposon mutagenesis and immunomagnetic separation (Lehti et al., 2013).
Keywords: Protein expression, Screening, Colony, Blotting, Agar plate
Materials and Reagents
Equipment
Procedure
Recipes
Acknowledgments
This protocol was modified from Sambrook, J. and Russell, D. (2001) Molecular Cloning: A Laboratory Manual (Cold Spring Harbor, NY: Cold Spring Harbor Laboratory). The authors would like to acknowledge University of Helsinki, Viikki Graduate School in Biosciences, the Academy of Finland (ERA-NET PathoGenoMics grant number 118982) and the European Network of Excellence in EuroPathoGenomics EPG (CEE LSHB-CT-2005-512061) for financial support.
References
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