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Intracellular protein determination using droplet-based immunoassays

Martino, Chiara and Zagnoni, Michele and Sandison, Mairi and Chanasakulniyom, Mayuree and Pitt, Andrew and Cooper, Jonathan (2011) Intracellular protein determination using droplet-based immunoassays. Analytical Chemistry, 83 (13). 5361–5368. ISSN 0003-2700

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Abstract

This paper describes the implementation of a sensitive, on-chip immunoassay for the analysis of intracellular proteins, developed using microdroplet technology. The system offers a number of analytical functionalities, enabling the lysis of low cell numbers, as well as protein detection and quantification, integrated within a single process flow. Cells were introduced into the device in suspension and were electrically lysed in situ. The cell lysate was subsequently encapsulated together with antibody-functionalized beads into stable, water-in-oil droplets, which were stored on-chip. The binding of intracellular proteins to the beads was monitored fluorescently. By analyzing many individual droplets and quantifying the data obtained against standard additions, we measured the level of two intracellular proteins, namely, HRas-mCitrine, expressed within HEK-293 cells, and actin-EGFP, expressed within MCF-7 cells. We determined the concentrations of these proteins over 5 orders of magnitude, from 50 pM to 1 μM. The results from this semiautomated method were compared to those for determinations made using Western blots, and were found not only to be faster, but required a smaller number of cells.

Item type: Article
ID code: 33143
Keywords: intracellular proteins, immunoassays, Electrical engineering. Electronics Nuclear engineering, Physical and theoretical chemistry, Analytical Chemistry
Subjects: Technology > Electrical engineering. Electronics Nuclear engineering
Science > Chemistry > Physical and theoretical chemistry
Department: Faculty of Engineering > Electronic and Electrical Engineering
Technology and Innovation Centre > Advanced Science and Technology
Technology and Innovation Centre > Bionanotechnology
Related URLs:
    Depositing user: Pure Administrator
    Date Deposited: 06 Sep 2011 16:46
    Last modified: 01 Aug 2014 05:06
    URI: http://strathprints.strath.ac.uk/id/eprint/33143

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