title: Expanding the available assays: adapting and validating In-Cell Westerns in microfluidic devices for cell-based assays.
abstract: Microfluidic methods for cellular studies can significantly reduce costs due to reduced reagent and biological specimen requirements compared with many traditional culture techniques. However, current types of readouts are limited and this lack of suitable readouts for microfluidic cultures has significantly hindered the application of microfluidics for cell-based assays. The In-Cell Western (ICW) technique uses quantitative immunocytochemistry and a laser scanner to provide an in situ measure of protein quantities in cells grown in microfluidic channels of arbitrary geometries. The use of ICWs in microfluidic channels was validated by a detailed comparison with current macroscale methods and shown to have excellent correlation. Transforming growth factor-β-induced epithelial-to-mesenchymal transition of an epithelial cell line was used as an example for further validation of the technique as a readout for soluble-factor-based assays performed in high-throughput microfluidic channels. The use of passive pumping for sample delivery and laser scanning for analysis opens the door to high-throughput quantitative microfluidic cell-based assays that integrate seamlessly with existing high-throughput infrastructure.
authors.affiliation: Department of Biomedical Engineering, Wisconsin Institutes for Medical Research, University of Wisconsin-Madison, Madison, Wisconsin 53726, USA.
authors.forename: Amy L
authors.initials: AL
authors.lastname: Paguirigan
authors.forename: John P
authors.initials: JP
authors.lastname: Puccinelli
authors.forename: Xiaojing
authors.initials: X
authors.lastname: Su
authors.forename: David J
authors.initials: DJ
authors.lastname: Beebe
mesh_terms: D000906:Antibodies / Q000276:immunology; D015153:Blotting, Western; D018929:Cell Culture Techniques; D002460:Cell Line; D058750:Epithelial-Mesenchymal Transition; D057166:High-Throughput Screening Assays; D006801:Humans; D007091:Image Processing, Computer-Assisted; D007118:Immunoassay; D046210:Microfluidic Analytical Techniques; D015203:Reproducibility of Results; D016212:Transforming Growth Factor beta / Q000494:pharmacology
chemical_list: D000906:Antibodies; D016212:Transforming Growth Factor beta