Use of a fluorescent stain for evaluating in vitro infection with Leishmania panamensis

Leishmaniasis is a group of endemic diseases produced by infection with Leishmania parasites and affects tropical and subtropical regions of the world. Due to the severe problems related to the treatment of this condition (resistance and toxicity), further studies are needed to evaluate new antileishmanial compounds. The activity of antileishmanial prototypes should be analyzed in models that allow a better interpretation of the findings with respect to natural infection. In this sense, the use of an infection model with macrophages and dendritic cells is a better than using promastigotes alone, in order to establish the potential leishmanicidal activity of a prototype compound. For infection analysis, staining with polychromatic dyes such as Giemsa plus microscopic examination is the gold standard. However, it is common to find problems associated with color uniformity, expertise of the observer, sensitivity and specificity of the technique. For this reason, it's necessary to develop tools and protocols to overcome such limitations. This study assessed the utility of the SYBR® Safe fluorescent dye, considering its affinity for nucleic acids as a useful property for staining the nucleus and kinetoplast of Leishmania parasites within an infected cell. Infection (and subsequent treatment) assays were performed in dendritic cells and macrophages infected with Leishmania panamensis parasites to compare SYBR® Safe and Giemsa stain for the same assay. Correlation coefficients were found to be above 0.9 for both techniques; however, unlike Giemsa, SYBR® Safe staining was easier and provided a clearer observation of internalized parasites. These results support the use of SYBR® Safe as a promising tool for evaluating potential antileishmanials given its advantages over the traditional technique.