Journal: Memórias do Instituto Oswaldo Cruz

Article Title: Past and future of trypanosomatids high-throughput phenotypic screening

doi: 10.1590/0074-02760210402

Figure Lengend Snippet: HCS-based assays employed in trypanosomatid drug discovery

Article Snippet: , L. donovani , Laboratory strain: MHOM/SD/62/1S-CL2D , Intracellular amastigotes , None , Human acute monocytic leukemia cells (THP-1) , Number of host cells (cytotoxicity evaluation), number of amastigotes per cell and infection ratio , Parasites DNA spots as well as cell nucleus and cytoplasm detected by Draq5 , 1,742 bioactive compounds from MedChem Express , Primary screening and cytotoxicity evaluation performed in the same assay. A similar method was used to generate the dose-response curves. Protocol based on a previous report (82) , (95).

Techniques: Infection, Immunostaining, Cytotoxicity Assay, Activity Assay, Luciferase, Proliferation Assay, Imaging, Time-Kill Assay, Derivative Assay, Expressing, Drug discovery, Staining, Labeling, SYBR Green Assay

Journal: Molecular Therapy. Methods & Clinical Development

Article Title: Accurate Titration of Infectious AAV Particles Requires Measurement of Biologically Active Vector Genomes and Suitable Controls

doi: 10.1016/j.omtm.2018.07.004

Figure Lengend Snippet: Immunofluorescence Analysis of Intracellular Localization of AAV8 Particles (A) Representative pictures of infected HeLa cells. Cells were non-infected (no AAV) or infected with AAV8 control or AAV8ΔVP1 at a multiplicity of 20,000 VG/cell, and then they were fixed after 1, 5, or 16 hr. Cell nuclei stained with DraQ5 appear in red, and assembled AAV8 particles stained with Alexa Fluor 555 appear in blue, green, or cyan, depending on their localization (cytoplasmic, intranuclear, or perinuclear, respectively). (B) Quantitative analysis of the immunofluorescence pictures. AAV8-assembled particles were quantified in the intranuclear, perinuclear, and cytoplasmic cellular compartments at 1, 5, and 16 hr post-infection with AAV8-GFP (upper panel) or AAV8ΔVP1-GFP (lower panel). Results obtained with AAV8ΔVP1 and AAV8 were compared by a two-tailed Mann-Whitney test for each cell compartment. *p < 0.05, **p < 0.005, ***p ≤ 0.0001; N = total number of AAV8 particles counted at each time point. Data are presented as mean ± SD.

Article Snippet: Cells were then incubated with anti-mouse Alexa Fluor 555 secondary antibody (1:200 in PBS), washed with PBS, and incubated with DraQ5 (1:1,000 in PBS) for nuclei staining (Interchim, Montluçon, France).

Techniques: Immunofluorescence, Infection, Control, Staining, Two Tailed Test, MANN-WHITNEY