Quantitative High-Throughput Assays for Flagella-Based Motility in Chlamydomonas Using Plate-Well Image Analysis and Transmission Correlation Spectroscopy

Abstract

Cilia are motile and sensory organelles with important roles in human development, physiology, and disease. Genetic defects in cilia produce a host of disease symptoms, including polycystic kidney disease, hydrocephalus, retinal degeneration, chronic bronchiectasis, infertility, and polydactyly. Currently, there are no known drugs for pharmacological remediation of ciliary defects. Small-molecule modulators of ciliary assembly or function would provide potential lead compounds for drug discovery efforts and would immediately be invaluable tools for a chemical biology approach to studying cilia. Here the author describes 2 assays for ciliary motility that are quantitative, automatable, cost-effective, and simple to implement. Both assays exploit cell-based strategies using the model organism Chlamydomonas reinhardtii. The first assay scores cilia-dependent gravitaxis by analyzing the cell distribution in wells of U-bottom microplates, using a simple and robust image analysis algorithm. The second assay measures motility directly by estimating the time required for cells to swim across a small illuminated aperture using a method equivalent to fluorescence correlation spectroscopy adapted to transmitted-light microscopy. The 2 assays have different advantages in terms of speed and sensitivity to small reductions in motility and may be most efficiently used in combination. (Journal of Biomolecular Screening 2009:133-141)

Keywords

ciliogenesis gravitaxis intraflagellar transport dynein inhibitor

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