Monitoring of the heat-shock response in Escherichia coli using an optical biosensor
2003 (English)In: Analytical Biochemistry, ISSN 0003-2697, Vol. 322, no 2, 156-163 p.Article in journal (Refereed) Published
A surface plasmon resonance (SPR) method for monitoring the concentration of the chaperone DnaK and its relation to physiological stress response in a recombinant Escherichia coli strain subjected to heat shock is described. The DnaK protein, an abundantly occurring representative of the heat-shock proteins, was used as a marker of physiological stress. The SPR biosensor instrument was used for label-free immunoaffinity detection directly in cell culture lysates using an anti-DnaK monoclonal IgG antibody immobilized on the sensor surface. The SPR method provides a fast response (<8min) and a reproducible (RSD<2%), accurate (comparison to the direct enzyme-linked immunosorbent assay), and sensitive (LOD<1nM) assay for determination of the DnaK level in cell culture lysates. The operational stability of the method was high compared to that of other SPR assays, the sensitivity decreased at only 2.7%/h. This allowed measurement of more than 220 samples per sensor surface. Storage stability was determined at 25°C (100% after 17h) and 10°C (101% after 1month). The method was validated by standard additions of DnaK (30, 60, and 120nM) with recovery indices in the range 95.7-103.7%. © 2003 Elsevier Inc. All rights reserved.
Place, publisher, year, edition, pages
2003. Vol. 322, no 2, 156-163 p.
DnaK analysis, Escherichia coli, Heat shock response analysis, Stress response sensor, Surface plasmon resonance
Engineering and Technology
IdentifiersURN: urn:nbn:se:liu:diva-46435DOI: 10.1016/j.ab.2003.07.019OAI: oai:DiVA.org:liu-46435DiVA: diva2:267331