A simple and rapid nonviral approach to efficiently transfect primary tissue-derived cells using polyethylenimine.

Abstract:

:This protocol outlines steps for optimizing the transfection of adherent primary mammalian cells using the readily available off-the-shelf cationic polymer, 25-kDa branched polyethylenimine (bPEI25). Transfection efficiency of cationic polymers varies among cell lines and is highly dependent on the conditions and environment in which complexes are formed. Factors requiring optimization include the salt concentration, volume, incubation time, mixing order and ratio of polymer to DNA. In this transfection protocol, complexes are prepared in 30 min, with analysis 24 h later; thus, experiments can be completed in 2 d. In this protocol, as an example, we describe the parameters we have optimized for the transfection of bone marrow stromal cells and normal human foreskin fibroblasts. By using this protocol, we have obtained transfection efficiencies comparable to lipofection. An appropriately optimized protocol enhances the utility of cationic polymers in transfecting mammalian cells, thereby providing an effective alternative to expensive commercial reagents.

journal_name

Nat Protoc

journal_title

Nature protocols

authors

Hsu CY,Uludağ H

doi

10.1038/nprot.2012.038

subject

Has Abstract

pub_date

2012-04-19 00:00:00

pages

935-45

issue

5

eissn

1754-2189

issn

1750-2799

pii

nprot.2012.038

journal_volume

7

pub_type

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