Are A549 cells easy to transfect?
It´s very easy. Invitrogen have protocols to how to transfect all cells, including A549. At least on these protocols have the plates and volume to applicate.
How much plasmid do you need for transfection?
Optimal amount of Universal Transfection Reagent used depends on cell type and is generally 1 – 3 µL per ug of plasmid DNA.
Can Lipofectamine be used in vivo?
As a prelude to direct in vivo gene transfer studies, we have compared the ability of two commercially available cationic lipids (Lipofectamine and DOSPER) to transfect C2C12 cells, a mouse muscle cell line in vitro. In addition, formu- lations suitable for in vivo injection were determined.
Is Lipofectamine toxic to cells?
Although Lipofectin showed the lowest toxicity to HepG2 cells (89.54% viability), it displayed the lowest transfection efficacy too (8.29%). Generally, HepG2 cells displayed resistance to the toxicity of majority of the reagents tested, with Lipofectamine 3000 (70.59 % viability), being the most toxic reagent.
How long does it take A549 cells to adhere?
A549 Cell Subculture Protocol A549 cells are cultured in complete media consisting of Dulbecco’s MEM modified with 10% FBS. The cells can be grown as adherent or in suspension in vitro. The A549 cell line grows easily and cell count doubling time is typically 24-40 hours.
What are A549 cells used for?
The A549 cells have been used to model the alveolar Type II pulmonary epithelium. Studies have shown that this can be particularly useful in research for studying the metabolic processing of lung tissue and for identifying mechanisms of drug delivery to the tissue.
How do you increase lipofectamine transfection efficiency?
Here are some tips that may help you improve your transfection success.
- Transfect healthy, actively dividing cells at a consistent cell density.
- Transfect using high-quality DNA.
- Optimize the amount of DNA used to transfect cells.
- Optimize the transfection reagent:DNA ratio.
How much plasmid do I need for electroporation?
The plasmid DNA amount is one of the parameters having significant transfection efficiency. The standard volume of injection is 50 μL, although depending on the muscle, the volumes vary from 10–100 μL (the smaller volume, the easier it is absorbed). For muscle, the DNA concentration should be 0.5–1.0 μg/μL.
How do lipid based transfection reagents work?
Lipid based reagents use cell membrane semi-permeability to deliver foreign nucleic acid molecules into a cell. Typically, the liposome is endocytosed by the cell and the nucleic acid cargo is released into the cytoplasm.
Which transfection reagent is best?
Under the tested conditions, ViaFect™ Reagent offered the best combination of transfection efficiency and low toxicity for most cell lines, making it an ideal choice when beginning transfection experiments with a new cell line.
How do you transfect A549 cells with Lipofectamine?
Transfecting A549 Cells. For each well of cells, dilute 1.5-2.5 μl of Lipofectamine® LTX into the above diluted DNA solution, mix gently and incubate for 25 minutes at room temperature to form DNA-Lipofectamine® LTX complexes. Remove growth medium from cells and replace with 0.5 ml of complete growth medium.
What is the protocol for Lipofectamine 2000?
Lipofectamine®2000 DNA Transfection Reagent Protocol Transfect cells according to the following chart. Volumes are given on a per-well basis. Each reaction mix is sufficient for triplicate (96-well), duplicate (24-well), and single well (6-well) transfections, and accounts for pipetting variations.
How do I transfect plasmid DNA into A549 cells?
Use this procedure to transfect plasmid DNA into A549 cells in a 24-well format (for other formats, see Scaling Up or Down Transfections, below). All amounts and volumes are given on a per well basis. The day before transfection, trypsinize and count the cells. Plate 4 x 10 4 cells per well in 0.5 ml of complete growth medium.
How to transfect A549 cells in tissue culture?
To transfect A549 cells in different tissue culture formats, vary the amounts of Lipofectamine® LTX Reagent, DNA, cells, medium and PLUS™ Reagent used in proportion to the relative surface area, as shown in the table (amounts given on a per well basis). 1 Surface areas may vary depending on the manufacturer.