1. Validation of quantitative real-time PCR for the in vitro assessment of antileishmanial drug activity.
- Author
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Gomes LI, Gonzaga FM, de Morais-Teixeira E, de Souza-Lima BS, Freire VV, and Rabello A
- Subjects
- Amphotericin B pharmacology, Animals, Cells, Cultured, DNA, Protozoan analysis, Dose-Response Relationship, Drug, Inhibitory Concentration 50, Leishmania infantum genetics, Mice, Mice, Inbred BALB C, Antiprotozoal Agents pharmacology, Leishmania infantum drug effects, Macrophages, Peritoneal parasitology, Real-Time Polymerase Chain Reaction standards
- Abstract
In vitro assays play an important role in the discovery and development of new antileishmanial drugs. The classic macrophage-amastigote models using murine peritoneal macrophages or human-monocyte derived macrophages as host cells are useful for drug screening. A major limitation of these models is the dependence on microscopic counting, a time-consuming and subjective method of analysis. The present study describes a detailed protocol for applying quantitative real-time PCR (qPCR) as an accurate and sensitive tool to assess parasite load in an amastigote-macrophage model. This assay can be performed in a standardized medium-to-high throughput procedure, replacing traditional readout of number of amastigote per macrophages by DNA load measurement., (Copyright © 2012 Elsevier Inc. All rights reserved.)
- Published
- 2012
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