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J Immunol Methods. 1998 Feb 1;211(1-2):79-86.

A novel method for DEAE-dextran mediated transfection of adherent primary cultured human macrophages.

Author information

1
Medicine and the AIDS Program, University of California San Francisco and San Francisco General Hospital, 94110, USA.

Abstract

Primary cultured human macrophages are difficult to transfect. We have developed a DEAE-dextran DNA transfection method that mediates the reproducible transfection of primary cultured adherent human macrophages. Three factors essential for successful transfection were identified: DEAE-dextran concentration, the quantity of DNA per transfection and the incubation time of the macrophages with the transfection medium. Maximum levels of luciferase expression were attained within 24 h and maintained for at least 56 h after transfection. While serum in the transfection medium attenuated transfection, the treatment of the macrophages with chloroquine, DMSO, or glycerol did not enhance transfection within this system. A CMV enhancer/promoter mediated substantially greater luciferase expression in the macrophages than either HIV or RSV LTRs. DEAE-dextran facilitated superior transfection compared to either cationic liposome and calcium phosphate methods, and was more practical compared to electroporation for multiple transfections. This transfection protocol provides a simple, inexpensive, reproducible method for the evaluation of gene expression in primary cultured adherent human macrophages.

PMID:
9617833
DOI:
10.1016/s0022-1759(97)00194-4
[Indexed for MEDLINE]

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