Improved protocol for efficient nonviral transfection of premature THP-1 macrophages.

Maess MB, Buers I, Robenek H, Lorkowski S.
Source: Nat Protocols
Publication Date: (2011)
Issue: 2011(5): 508-11
Research Area:
Cancer Research/Cell Biology
Immunotherapy / Hematology
Nucleofection of stimulated adherent THP-1 cells with human monocytes kit and program. Very good recovery with monocytes recovery medium and higher HSA concentration.
INTRODUCTION: The human monocytic leukemia cell line THP-1 is a widely used model for investigating monocyte and macrophage biology. Successful transfection of THP-1 monocytes with subsequent phorbol 12-myristate 13-acetate (PMA)-induced differentiation into macrophages is not a trivial matter, because according to previous transfection protocols, cell viability is lost almost completely within 24 h of PMA treatment following transfection. This protocol constitutes an optimized version of a previously published protocol by our group. It describes a procedure for transfecting premature THP-1 macrophages, which subsequently can be further differentiated into mature macrophages by PMA without a loss of cell viability. Transfection of THP-1 cells with plasmids or small interfering RNA (siRNA) is achieved by electroporation using the Lonza Nucleofector technology (Basel, Switzerland). This technique allows for the efficient nonviral delivery of plasmids, DNA, RNA, or siRNA into primary cells or cell lines even if the cells are not or are only slowly proliferating. Such cells are usually rather difficult to transfect by nonviral approaches. This means that only viral approaches would be left, which are expensive and labor-intensive and require laboratories complying with the respective safety regulations. The protocol described here is an efficient and convenient alternative.