Transfections allow for transient expression of a gene of interest in a target cell line and can be useful for short term studies of protein function. We specifically use this protocol with Lenti-X 293T cells, a cell line optimized for production of lentiviral vectors. Naprosen and plaquenil Acr hydroxychloroquine Plaquenil toxicity reversible Chloroquine-containing media were removed and the cells were washed with phosphate buffered saline before they were infected with SARS-CoV 0.5 multiplicity of infection for 1 h in the absence of chloroquine. Virus was then removed and the cells were maintained in Opti-MEM Invitrogen for 16–18 h in the absence of chloroquine. Chloroquine CQ is a lysosomotropic agent with an extensive range of biological effects 1. Historically known for its anti-malarial activity, chloroquine is a widely used biological research tool for studying autophagy inhibition. Research studies demonstrate that chloroquine accumulates in acidic lysosomes and increases the lysosomal pH. Chloroquine enters the red blood cell by simple diffusion, inhibiting the parasite cell and digestive vacuole. Chloroquine then becomes protonated to CQ2+, as the digestive vacuole is known to be acidic pH 4.7; chloroquine then cannot leave by diffusion. Last Upload: June 10, 2016 Day 0: Seed Lenti-X 293T cells (this cell line is optimized for production of lentiviral vectors) Day 1 (pm): Transfect Cells Day 2 (am): 18h post transfection - Remove media, replace with fresh media Day 3 or more (am): Observe fluorescence, harvest cells, or perform your experiment *Pro-Tips* Different brands and lots of FBS can promote or inhibit transfection. This approach can be adapted for different cell lines and different transfection reagents. Chloroquine invitrogen Chloroquine Indications, Side Effects, Warnings -, CST - Chloroquine Hydroxychloroquine and white spots on skinChloroquine spcChloroquine tablets side effects Invitrogen Anti-Rabbit IgG H+L Highly Cross-Adsorbed Secondary Antibody, Catalog # A-11034. Tested in Immunofluorescence IF, Immunocytochemistry ICC and Flow Cytometry Flow applications. This antibody is cross-adsorbed against bovine igg, goat igg, mouse igg, rat igg and human igg. Supplied as 1 mg purified secondary antibody 2 mg/mL. Goat anti-Rabbit IgG H+L Highly Cross-Adsorbed, Alexa Fluor® 488 A.. Chloroquine - Wikipedia. Chloroquine Diphosphate Salt, MP Biomedicals Fisher Scientific. Post-infection chloroquine treatment reduces SARS-CoV infection and spread. Vero E6 cells were seeded and infected as described for Fig. 1 except that chloroquine was added only after virus adsorption. Cells were maintained in Opti-MEM Invitrogen containing. Chloroquine is an antimalarial drug that has been used in humans for many years. In recent years, Chloroquine has been shown to inhibit autophagy and induce apoptosis in malignant cells and thus has been tested in various experimental model systems and in human clinical trials. Alexa Fluor R products are provided under an intellectual property license from Life Technologies Corporation. The purchase of this product conveys to the buyer the non-transferable right to use the purchased product and components of the product only in research conducted by the buyer whether the buyer is an academic or for-profit entity.