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Development of an Alamar Blue^TM Viability Assay in 384-Well Format for High Throughput Whole Cell Screening of Trypanosoma brucei brucei Bloodstream Form Strain 427

There is an urgent need for new compounds for the drug development pipeline for treatment of patients with African sleeping sickness. One approach for identifying such compounds is by high throughput screening (HTS) of compound collections. For time and cost considerations, there is a need for the development of an assay that uses at least 384-well formats. To our knowledge, there are currently no viability assays for whole cell screening of trypanosomes in the 384-well plate format. We have developed and optimized an Alamar Blue viability assay in a 384-well format for Trypanosoma brucei brucei bloodstream form strain 427 (BS427). The assay had a Z' > 0.5 and tolerated a final dimethyl-sulfoxide concentration of 0.42%. Drug sensitivity was compared with those reported from previously developed 96-well methods and was found to be comparable. The sensitivity and cost benefit of the Alamar Blue assay make it an excellent candidate for HTS application.

Received January 12, 2009. Accepted for publication July 7, 2009.

Acknowledgments: We thank Dr. Achim Schnaufer (University of Edinburgh, Edinburgh, Scotland) who while at the Seattle Biomedical Research Institute, kindly supplied the trypanosome cell stock used throughout this study. We also thank Gillian Fisher for assistance with studies to evaluate the effects of incubation time with Alamar Blue.

Financial support: This study was supported by the Drugs for Neglected Diseases Initiative.

^* Address correspondence to Vicky M. Avery, Eskitis Institute for Cell and Molecular Therapies, Griffith University, Eskitis Building N27, Brisbane Innovation Park, Don Young Road, Nathan, Queensland 4111, Australia. E-mail: v.avery{at}griffith.edu.au

Authors’ address: Melissa L. Sykes and Vicky M. Avery, Eskitis Institute for Cell and Molecular Therapies, Griffith University, Eskitis Building N27, Brisbane Innovation Park, Don Young Road, Nathan, Queensland 4111, Australia, E-mails: m.sykes{at}griffith.edu.au and v.avery{at}griffith.edu.au.