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. Which autoimmune not treat with plaquenil Plaquenil retinal testing guidelines Chloroquine diphosphate 52 mg Deionized, distilled H 2 O 1 mL Dissolve 52 mg of chloroquine diphosphate in 1 mL of deionized, distilled H 2 O. Sterilize the solution by passing it through a 0.22-µm filter. Store the filtrate in foil-wrapped tubes at −20°C. Transfection Procedure. The morning of the transfection day, replace the media with fresh DMEM without PSG and containing 10 uL of 25 mM chloroquine optional. Wait ~5h before going onto the next step. Prepare DNA in a sterile 1.5 mL tube or tubes as needed. Use 100 uL per well. Add 3 uL PEI per ug of DNA in OptiMEM. A more recent usage is for DNA transfection.7,8 When used at 100 µM, it intercalates into DNA, increasing tranfection efficiency.9 Precautions and Disclaimer For Laboratory Use Only. Not for drug, household or other uses. Preparation Instructions This product is soluble in water 50 mg/ml. A 10% solution in water has a pH of 3.5-4.5. A 7.15%. 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 dna transfection Chloroquine diphosphate salt powder or crystals, 98.5-101.0%., PEI Mediated Plasmid Transfection - Bridges Lab Protocols Avloclor 250mg chloroquine phosphateOct appearance of plaquenil toxicityPlaquenil retionpathyEye exam when taking plaquenil A pH-responsive peptide fragment modelled on the influenza virus haemagglutinin INF7-SGSC can promote the transfectional activity of polyL-lysine pLL/DNA complexes against 293 cells. Chloroquine and amphipathic peptide helices show synergistic.. Chloroquine diphosphate salt - Sigma-Aldrich. DEAE-Dextran Transfection of Lymphocyte Cell Lines. Transfection-mediated toxicity usually leads to changes in gene expression, even in the absence of any visual toxicity. Therefore, it is important to choose a transfection method that can mitigate both observed as well as non-apparent cellular toxicity. Cationic lipids and polymers also form complexes with DNA through electrostatic interactions. Chloroquine has been used to increase transfection efficiency for calcium phosphate, DEAE-dextran, and some liposome-mediated gene delivery 63,65–68. However, effects of lysosomotropic agents on polymer and some lipid based methods of gene delivery are still controversial in the literature. Add 22 l of 25mM chloroquine to each 15cm dish for 10cm plate 7.8 l of chloroquinei.e. final concentration = 25 M. The cells should be transfected within 5-10’ of adding the chloroquine. 7. For 15cm plates, add 1.69ml of the transfection buffer to the DNA-CaCl2 solution by “bubbling”.