title: Digital holographic microscopy: a quantitative label-free microscopy technique for phenotypic screening.
abstract: Digital Holographic Microscopy (DHM) is a label-free imaging technique allowing visualization of transparent cells with classical imaging cell culture plates. The quantitative DHM phase contrast image provided is related both to the intracellular refractive index and to cell thickness. DHM is able to distinguish cellular morphological changes on two representative cell lines (HeLa and H9c2) when treated with doxorubicin and chloroquine, two cytotoxic compounds yielding distinct phenotypes. We analyzed parameters linked to cell morphology and to the intracellular content in endpoint measurements and further investigated them with timelapse recording. The results obtained by DHM were compared with other optical label-free microscopy techniques, namely Phase Contrast, Differential Interference Contrast and Transport of Intensity Equation (reconstructed from three bright-field images). For comparative purposes, images were acquired in a common 96-well plate format on the different motorized microscopes. In contrast to the other microscopies assayed, images generated with DHM can be easily quantified using a simple automatized on-the-fly analysis method for discriminating the different phenotypes generated in each cell line. The DHM technology is suitable for the development of robust and unbiased image-based assays.
authors.forename: Benjamin
authors.initials: B
authors.lastname: Rappaz
authors.forename: Billy
authors.initials: B
authors.lastname: Breton
authors.forename: Etienne
authors.initials: E
authors.lastname: Shaffer
authors.affiliation: Biomolecular Screening Facility, Ecole Polytechnique Fédérale de Lausanne (EPFL), Station 15, Lausanne 1015, Switzerland. gerardo.turcatti@epfl.ch.
authors.forename: Gerardo
authors.initials: G
authors.lastname: Turcatti
mesh_terms: D000818:Animals; D001681:Biological Assay; D002460:Cell Line; D002738:Chloroquine / Q000494:pharmacology; D003603:Cytotoxins / Q000494:pharmacology; D004317:Doxorubicin / Q000494:pharmacology; D006367:HeLa Cells; D006696:Holography / Q000379:methods*; D006801:Humans; D007091:Image Processing, Computer-Assisted; D008858:Microscopy, Phase-Contrast / Q000379:methods*; D032383:Myocytes, Cardiac / Q000187:drug effects / Q000648:ultrastructure*; D010641:Phenotype; D051381:Rats; D059008:Time-Lapse Imaging
chemical_list: D003603:Cytotoxins; D004317:Doxorubicin; D002738:Chloroquine