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Background: Fully differentiated adipocytes are considered to be refractory to introduction of siRNA via lipid-based transfection. However, large scale siRNA-based loss-of-function screening of adipocytes using either electroporation or virally-mediated transfection approaches can be prohibitively complex and expensive. Methodology/Principal Findings: We present a method for introducing small interfering RNA (siRNA) into differentiated 3T3-L1 adipocytes and primary human adipocytes using an approach based on forming the siRNA/ cell complex with the adipocytes in suspension rather than as an adherent monolayer, a variation of "reverse transfection". Conclusions/Significance: Transfection of adipocytes with siRNA by this method is economical, highly efficient, has a simple workflow, and allows standardization of the ratio of siRNA/cell number, making this approach well-suited for high-throughput screening of fully differentiated adipocytes. © 2009 Kilroy et al.

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