A parasite-rescue and transformation assay with THP1 cells infected in vitro with Leishmania donovani has been optimized for anti-leishmanial drug screening. The assay involves differentiation of THP1 cells, infection with promastigotes, treatment with test drugs, controlled lysis of the infected macrophages, rescue of amastigotes, transformation to promastigotes and monitoring promastigote growth and proliferation with a fluorometric assay.
This video article details a straightforward in vivo methodology that can be used to systematically and efficiently characterize components of complex signaling pathways and regulatory networks in many invertebrate embryos.
Methods to prepare and characterize the physicochemical properties and bioactivity of neutrally-charged, pH-responsive siRNA nanoparticles are presented. Criteria for successful siRNA nanomedicines such as size, morphology, surface charge, siRNA loading, and gene silencing are discussed.
We present a cost-effective method to the scratch migration assay that provides a new approach for determining cell migration without the use of equipment-intensive methods. While fibroblasts were used in this protocol, it can be adapted and utilized to study additional cell types and influences on cell migration.
Fast photochemical oxidation of proteins is an emerging technique for the structural characterization of proteins. Different solvent additives and ligands have varied hydroxyl radical scavenging properties. To compare the protein structure in different conditions, real-time compensation of hydroxyl radicals generated in the reaction is required to normalize reaction conditions.
This protocol presents a method to use inline radical dosimetry and a plasma light source to perform flash oxidation protein footprinting. This method replaces the hazardous UV laser to simplify and improve the reproducibility of fast photochemical oxidation of protein studies.
Formation of actomyosin bundles in vitro and measuring myosin ensemble force generation using optical tweezers is presented and discussed.
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