Preparation and Characterization of A Nanoliposomal Vaccine of pcLACK Candidate Against Cutaneous Leishmaniasis.

BACKGROUND: Leishmaniasis is a public health problem and endemic in countries of the tropics and subtropics. An ongoing project with naked LACK (Leishmania homolog of receptors for activated C-kinase) demonstrated that this case of the gene is entirely susceptible to immune response and it does enter the cells effectively. This study aimed at developing a procedure to prepare a type of lipid nanoparticles overloaded with plasmid LACK (pcLACK) for usage as Leishmania major (L. major) nanoliposomal vaccine. MATERIALS AND METHODS: The single-gene expression plasmid of pcLACK was encoded in the LACK antigen. Nanoparticles were set up by thin film procedure using cationic lipids 1, 2-Dioleoyl- 3-Trimethylammonium propane (DOTAP), 1, 2-Dioleoyl-snGlycero-3-Phosphoethanolamine (DOPE), and cholesterol in a molar proportion of 2:1:1 molar ratio. Using dynamic light scattering, the particle diameters of empty and loaded lipoplexes were measured in triplicate. The zeta-potential (ζ) was measured with the same instrument using the zeta potential mode as the average of 20 measurements by diluting the particles into a low salt buffer. RESULTS: The results of the sustainability studies of Liposome-pcLACK formulation showed that there were no significant physical changes up to the 30th day of stability study at the storage condition of 4°C. However, there were significant changes in the formulation content during storage at 25°C for 30 days (204.2±0.90 at Day 30 compared with 207.2±0.26 nm at Day 0). It was observed that the prepared nanoliposomal formulation had more stability under refrigeration. CONCLUSION: Immunostimulatory cationic lipids bearing a pcLACK encapsulation could serve as an effective delivery system.

A nanodiagnostic colorimetric assay for 18S rRNA of Leishmania pathogens using nucleic acid sequence-based amplification and gold nanorods.

BACKGROUND AND OBJECTIVE: We describe here a nanodiagnostic colorimetric assay for 18S rRNA of Leishmania pathogens that uses nucleic acid sequence-based amplification (NASBA) and gold nanorods (GNRs). METHODS: NASBA primers targeting 18S rRNA were used for amplification of RNA in an isothermal process. The electrostatic interactions between the phosphate groups of the RNA amplicons and the cetyl trimethylammonium bromide layer on the GNRs resulting in their aggregation. This phenomenon changes the color of the GNR solution from red to purple. RESULTS: Our data showed 100% sensitivity and 80% specificity with the colorimetric assay compared with results using reverse transcription polymerase chain reaction. CONCLUSION: The nanodiagnostic method we describe simplifies the detection of NASBA amplicons without the need for gel electrophoresis.