title: Digital microfluidic immunocytochemistry in single cells.
abstract: We report a new technique called Digital microfluidic Immunocytochemistry in Single Cells (DISC). DISC automates protocols for cell culture, stimulation and immunocytochemistry, enabling the interrogation of protein phosphorylation on pulsing with stimulus for as little as 3 s. DISC was used to probe the phosphorylation states of platelet-derived growth factor receptor (PDGFR) and the downstream signalling protein, Akt, to evaluate concentration- and time-dependent effects of stimulation. The high time resolution of the technique allowed for surprising new observations-for example, a 10 s pulse stimulus of a low concentration of PDGF is sufficient to cause >30% of adherent fibroblasts to commit to Akt activation. With the ability to quantitatively probe signalling events with high time resolution at the single-cell level, we propose that DISC may be an important new technique for a wide range of applications, especially for screening signalling responses of a heterogeneous cell population.
authors.affiliation: Institute of Biomaterials and Biomedical Engineering, University of Toronto, 164 College Street, Toronto, Ontario M5S 3G9, Canada.|Donnelly Centre for Cellular and Biomolecular Research, 160 College Street, Toronto, Ontario M5S 3E1, Canada.
authors.forename: Alphonsus H C
authors.initials: AHC
authors.lastname: Ng
authors.affiliation: Institute of Biomaterials and Biomedical Engineering, University of Toronto, 164 College Street, Toronto, Ontario M5S 3G9, Canada.|Donnelly Centre for Cellular and Biomolecular Research, 160 College Street, Toronto, Ontario M5S 3E1, Canada.|Department of Chemical Engineering and Applied Chemistry, 200 College Street, Toronto, Ontario M5S 3E5, Canada.
authors.forename: M Dean
authors.initials: MD
authors.lastname: Chamberlain
authors.affiliation: Institute of Biomaterials and Biomedical Engineering, University of Toronto, 164 College Street, Toronto, Ontario M5S 3G9, Canada.|Donnelly Centre for Cellular and Biomolecular Research, 160 College Street, Toronto, Ontario M5S 3E1, Canada.
authors.forename: Haozhong
authors.initials: H
authors.lastname: Situ
authors.affiliation: Donnelly Centre for Cellular and Biomolecular Research, 160 College Street, Toronto, Ontario M5S 3E1, Canada.
authors.forename: Victor
authors.initials: V
authors.lastname: Lee
authors.affiliation: Institute of Biomaterials and Biomedical Engineering, University of Toronto, 164 College Street, Toronto, Ontario M5S 3G9, Canada.|Donnelly Centre for Cellular and Biomolecular Research, 160 College Street, Toronto, Ontario M5S 3E1, Canada.|Department of Chemistry, University of Toronto, 80 Saint George Street, Toronto, Ontario M5S 3H6, Canada.
authors.forename: Aaron R
authors.initials: AR
authors.lastname: Wheeler
mesh_terms: D000818:Animals; D018929:Cell Culture Techniques; D002460:Cell Line; D005347:Fibroblasts / Q000378:metabolism*; D006801:Humans; D007150:Immunohistochemistry / Q000295:instrumentation / Q000379:methods*; D061986:MCF-7 Cells; D051379:Mice; D046210:Microfluidic Analytical Techniques / Q000295:instrumentation / Q000379:methods*; D044085:Microfluidics; D041681:NIH 3T3 Cells; D010766:Phosphorylation; D010982:Platelet-Derived Growth Factor / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism*; D015398:Signal Transduction
chemical_list: D010982:Platelet-Derived Growth Factor; D051057:Proto-Oncogene Proteins c-akt