Real-Time PCR Assay for detection and quantification of Leishmania: standardization, positive control, validation, and intra-laboratory assay

His study aimed to develop a method to investigate PCR sensitivity for diagnosis and ensure reproducibility for parasite load quantification in tissues based on qPCR. In the first step, genes were selected to quantify the parasite load; then, a standard was developed to quantify the concentration of different Leishmania species. These tools were evaluated in intra-laboratory assays. The sensitivity was determined as 0.01 parasites/μL, and the method was reproducible with 100% concordance among human participants in the intra-laboratory validation study. Furthermore, the results demonstrated the specificity of the method in detecting the genus Leishmania without showing cross-reaction with Trypanosoma cruzi or human DNA.