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  • Development of a sandwich ELISA to detect Leishmania 40S ribosomal protein S12 antigen from blood samples of visceral leishmaniasis patients.

Development of a sandwich ELISA to detect Leishmania 40S ribosomal protein S12 antigen from blood samples of visceral leishmaniasis patients.

BMC infectious diseases (2018-10-05)
Wen-Wei Zhang, Ayan Kumar Ghosh, Raodoh Mohamath, Jacqueline Whittle, Alessandro Picone, Patrick Lypaczewski, Momar Ndao, Randall F Howard, Pradeep Das, Steven G Reed, Greg Matlashewski
ABSTRACT

Visceral leishmaniasis (VL), caused by Leishmania donovani complex parasites, is a neglected parasitic disease that is generally fatal if untreated. Despite decades of research to develop a sensitive VL diagnostic test, definitive diagnosis of VL still mainly relies on the visualization of the parasite in aspirates from the spleen, liver or bone marrow, an invasive and dangerous process with variable sensitivity. A sensitive assay that can detect Leishmania antigen from blood samples will help confirm cause, cure or recurrence of VL. In this study, rabbit polyclonal antibodies were raised against eight recombinant Leishmania proteins that are highly abundant in Leishmania. The antibodies were purified and labeled with biotin for developing a prototype sandwich enzyme-linked immunosorbent assay (ELISA). The ELISA for the Leishmania 40S ribosomal protein S12 detected target antigen with the highest sensitivity and specificity and could detect 1 pg of purified protein or as few as 60 L. donovani parasites. The 40S ribosomal protein S12 sandwich ELISA could detect the target antigen from Peripheral Blood Mononuclear Cell (PBMC) samples in 68% of VL patients and post-kala-azar dermal leishmaniasis (PKDL) patients, providing an estimation of parasitemia ranging from 15 to 80 amastigotes per ml of blood. These results indicate that the 40S ribosomal protein S12 sandwich ELISA warrants further tests with more clinical samples of VL patients and other parasitic diseases. It is hopeful that this ELISA could become a useful tool for confirming VL diagnosis, monitoring treatment progress, disease recurrence and possibly detecting asymptomatic Leishmania infections with a high parasite load.

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Sigma-Aldrich
Medium 199, With Hanks′ salts and L-glutamine, without sodium bicarbonate, powder, suitable for cell culture