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High-throughput screening of amastigotes of Leishmania donovani clinical isolates against drugs using a colorimetric beta-lactamase assay.
Title | High-throughput screening of amastigotes of Leishmania donovani clinical isolates against drugs using a colorimetric beta-lactamase assay. |
Publication Type | Journal Article |
Year of Publication | 2009 |
Authors | Mandal, S, Maharjan, M, Ganguly, S, Chatterjee, M, Singh, S, Buckner, FS, Madhubala, R |
Journal | Indian J Exp Biol |
Volume | 47 |
Issue | 6 |
Pagination | 475-9 |
Date Published | 2009 Jun |
ISSN | 0019-5189 |
Keywords | Animals, Antiprotozoal Agents, beta-Lactamases, Biological Assay, Genes, Reporter, Humans, Leishmania donovani, Leishmaniasis, Cutaneous, Leishmaniasis, Visceral |
Abstract | A simple colorimetric beta-lactamase assay for quantifying Leishmania amastigotes in macrophages grown in microtiter plates has been reported. The beta-lactamase gene was integrated into the rRNA region of the genome, thereby allowing for high-level stable expression of the enzyme. Both visceral leishmaniasis (VL) and post-kala azar dermal leishmaniasis (PKDL) isolates were transfected with beta-Lactamase gene. These beta-lactamase-expressing promastigotes were used for infecting intracellular J774A.1 macrophages in vitro. Quantification was done by a colorimetric readout with CENTA beta-lactamase as substrate and with an optical density plate reader. The assay was carried out in 96-well plates. Results obtained demonstrate that this methodology could be a valuable high-throughput screening assay for checking efficacy of anti-leishmanial drugs in the clinical isolates. |
Alternate Journal | Indian J. Exp. Biol. |
PubMed ID | 19634714 |