Vishal Gaurav - Development of a live cell-based immunofluorescent assay for the detection of pharmaceutically-relevant secondary metabolites in plant cells

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  Publication Details (including relevant citation   information):

  Authors: Vishal Gaurav, Susan C. Roberts

  Publication date: 2009

  Conference name: The American Chemical Society (ACS) Proceedings

  Publisher: The ACS National Meeting, Salt Lake City, UT


  Paclitaxel, an important anti-cancer drug, is a complex   diterpenoid synthesized in Taxus plant cell cultures. To unravel   the inherent culture heterogeneity and variability in production,   our laboratory is investigating the regulation of cellular   metabolism in distinct cell populations, isolated using flow   cytometric cell sorting technologies. A novel indirect   immunofluorescent assay was developed to detect paclitaxel   accumulation in live plant cells to enable sorting of both high   and low accumulating populations. This assay utilized a mouse   monoclonal anti-paclitaxel antibody in conjunction with a   PEconjugated goat anti-mouse secondary antibody. Concomitantly,   cell viability was assessed using staining with fluorescein   diacetate to ensure that stained cells were undamaged and   remained intact following the procedure. Experiments were   performed with variations in antibody concentrations, wash steps   and incubation times to establish an optimal procedure.

  Address (URL): fluorescent_assay_for_the_detection_of_pharmaceutically-relevant_secondary_meta b olites_in_plant_cells.pdf