title: Improved visualization and quantitative analysis of drug effects using micropatterned cells.
abstract: To date, most HCA (High Content Analysis) studies are carried out with adherent cell lines grown on a homogenous substrate in tissue-culture treated micro-plates. Under these conditions, cells spread and divide in all directions resulting in an inherent variability in cell shape, morphology and behavior. The high cell-to-cell variance of the overall population impedes the success of HCA, especially for drug development. The ability of micropatterns to normalize the shape and internal polarity of every individual cell provides a tremendous opportunity for solving this critical bottleneck (1-2). To facilitate access and use of the micropatterning technology, CYTOO has developed a range of ready to use micropatterns, available in coverslip and microwell formats. In this video article, we provide detailed protocols of all the procedures from cell seeding on CYTOOchip micropatterns, drug treatment, fixation and staining to automated acquisition, automated image processing and final data analysis. With this example, we illustrate how micropatterns can facilitate cell-based assays. Alterations of the cell cytoskeleton are difficult to quantify in cells cultured on homogenous substrates, but culturing cells on micropatterns results in a reproducible organization of the actin meshwork due to systematic positioning of the cell adhesion contacts in every cell. Such normalization of the intracellular architecture allows quantification of even small effects on the actin cytoskeleton as demonstrated in these set of protocols using blebbistatin, an inhibitor of the actin-myosin interaction.
authors.affiliation: CYTOO Cell Architects, Grenoble, France. sdegot@cytoo.com
authors.forename: Sébastien
authors.initials: S
authors.lastname: Degot
authors.forename: Muriel
authors.initials: M
authors.lastname: Auzan
authors.forename: Violaine
authors.initials: V
authors.lastname: Chapuis
authors.forename: Anne
authors.initials: A
authors.lastname: Béghin
authors.forename: Amélie
authors.initials: A
authors.lastname: Chadeyras
authors.forename: Constantin
authors.initials: C
authors.lastname: Nelep
authors.forename: Maria Luisa
authors.initials: ML
authors.lastname: Calvo-Muñoz
authors.forename: Joanne
authors.initials: J
authors.lastname: Young
authors.forename: François
authors.initials: F
authors.lastname: Chatelain
authors.forename: Alexandra
authors.initials: A
authors.lastname: Fuchs
mesh_terms: D000199:Actins / Q000037:antagonists & inhibitors / Q000378:metabolism; D002448:Cell Adhesion; D003584:Cytological Techniques / Q000379:methods*; D003599:Cytoskeleton / Q000187:drug effects / Q000378:metabolism; D004353:Drug Evaluation, Preclinical / Q000379:methods*; D006367:HeLa Cells; D006576:Heterocyclic Compounds, 4 or More Rings / Q000494:pharmacology; D006801:Humans; D009218:Myosins / Q000037:antagonists & inhibitors / Q000378:metabolism; D013194:Staining and Labeling / Q000379:methods
chemical_list: D000199:Actins; D006576:Heterocyclic Compounds, 4 or More Rings; C472645:blebbistatin; D009218:Myosins