title: On-chip enzyme quantification of single Escherichia coli bacteria by immunoassay-based analysis.
abstract: Individual bacteria of an isogenic population can differ significantly in their phenotypic characteristics. This cellular heterogeneity is thought to increase the adaptivity to environmental changes on a population level. Analytical methods for single-bacteria analyses are essential to reveal the different factors that may contribute to this cellular heterogeneity, among them the stochastic gene expression, cell cycle stages and cell aging. Although promising concepts for the analysis of single mammalian cells based on microsystems technology were recently developed, platforms suitable for proteomic analyses of microbial cells are by far more challenging. Here, we present a microfluidic device optimized for the analysis of single Escherichia coli bacteria. Individual bacteria are captured in a trap and isolated in a volume of only 155 pL. In combination with an immunoassay-based analysis of the cell lysate, the platform allowed the selective and sensitive analysis of intracellular enzymes. The limit of detection of the developed protocol was found to be 200 enzymes. Using this platform, we could investigate the levels of β-galactosidase in cells grown under different nutrient conditions. We successfully determined the enzyme copy numbers in cells cultured in defined medium (3517 ± 1578) and in complex medium (4710 ± 2643), and verified the down-regulation of expression in medium that contained only glucose as carbon source. The strong variations we found for individual bacteria confirm the phenotype heterogeneity. The capability to quantify proteins and other molecules in single bacterial lysates is encouraging to use the new analysis platform in future proteomics studies of isogenic bacteria populations.
authors.affiliation: Department of Chemistry and Applied Biosciences, ETH Zurich , CH-8093 Zürich, Switzerland.
authors.forename: Simone
authors.initials: S
authors.lastname: Stratz
authors.forename: Klaus
authors.initials: K
authors.lastname: Eyer
authors.forename: Felix
authors.initials: F
authors.lastname: Kurth
authors.forename: Petra S
authors.initials: PS
authors.lastname: Dittrich
mesh_terms: D004926:Escherichia coli / Q000302:isolation & purification*; D007118:Immunoassay / Q000379:methods*; D056656:Lab-On-A-Chip Devices