Fast & Easy 3-Step Lipid Transfection Protocol

Product No. L3287

Introduction

Escort IV is a liposome suspension composed of a polycationic lipid and a neutral, non-transfecting lipid compound.  Transfection using liposomes is a commonly used method for the introduction of DNA into eukaryotic cells. This technique has been used to obtain both transient and stable transfections in a wide variety of cell types (see below).  The procedure is based on the formation of a complex between the plasmid DNA and the lipid reagent, which adheres to the cell surface and is taken up by the cell, presumably by endocytosis, releasing the DNA into the cytoplasm.

Precautions and Disclaimer

This product is for R&D use only, not for drug, household, or other uses. Please consult the Safety Data Sheet for information regarding hazards and safe handling practices.

Procedure for transfecting adherent primary cells and cell lines

The following protocol is designed for an experienced user to obtain good efficiency across a variety of cell types.  If you require a more detailed protocol, see the full version here. For highest transfection efficiency, use the Optimization Protocol to determine the best set of conditions for your cells.

Note: for best results, use a seeding density that will provide >70% confluence about 12 hours later.


STEP 1: Prepare Complexes

Dilute the plasmid DNA in medium in tube A.  In tube B, dilute the lipid reagent in medium. Add tube A to tube B and mix gently before incubating 15-30 minutes at room temperature.

  Tube A Tube B  
Culture Dish DNA Volume (µL) Medium volume (µL) Escort IV Transfection Reagent volume (µL) Medium volume (µL) Approximate volume of media during transfection
24 well plate 0.5 9.5 0.5 9.5 500 µL
12 well plate 1 49 1 49 1 mL
6 well plate or 3.5 cm dish 2 98 2 98 2 mL
10 cm dish 5 495 5 495 5 mL


STEP 2: Treat Cells

Remove half the medium from each cell culture dish and discard. Add complexes drop-wise to the cell cultures, and swirl the plates gently to distribute the complexes evenly over the cells. Incubate the cells 5-18 hours at 37°C.

STEP 3: Change Medium
Change the medium (use regular growth medium), and allow the cells in continue incubating 24 – 72 more hours, until you are ready to perform further analysis.