Reverse transfection

We may also find that Sabatini s reverse transfection method of creating "live cell" microarrays offers even greater advantage for drug discovery and development (Ziauddin and Sabatini, 2001). Their method, relying on arrayed cDNA expression vectors to transform adherent cells, provides localized real-time gene expression analysis of the putative gene product. 

Mishina YM, Wilson CJ, Bmett L, Smith JJ, Stoop-Myer C, Jong S, Amaral LP, Pedersen R, Lyman SK, Myer VE, Kreider BL, Thompson CM (2004) Multiplex GPCR assay in reverse transfection cell microarrays. J Biomol Screen 9 196—207... 

E. Palmer, and T. Freeman, Investigation into the use of C- and N-terminal GFP fusion proteins for sub-cellular localization studies using reverse transfection microarrays, Compar. Funct. Genom., 2004, 5(4), 342-353. 

Fig. 2 Cell arrays can be applied for screening of protein-protein interactions in mammalian cells. Expression plasmids containing cDNAs for bait, reporter, and prey proteins (A) are spotted on the glass surface (B) and reverse transfected into adhesive cell line (C). After 3 days of incubation, proteins interactions are detected by the measurement of the reporter protein fluorescence (D). Since transfected cell clusters are separated by nontransfected cells, hundreds of different prey constructs can be investigated in a single experiment...

Tissue engineering scaffold for DNA delivery by cationic polymers. Biomimetic scaffolds can be encapsulated with growth factors and MSCs are seeded onto their surface [top]. Polymeric release bottom left) consists in the entrapment of the complexes between cationic polymers and DNA within the biomaterial for release into the environment. Conversely, substrate-mediated delivery bottom right), also termed reverse transfection delivery, employs the immobilization of complexes to the biomaterial. MSCs can internalize the complexes either directiy or by degrading the linkage between the biomaterial and DNA complexes. 

Nguyen YT, Kim HK, Kwon JS et al (2010) Efficient transfer of reporter gene-loaded nanoparticles to Ixme marrow stromal cells (Dl) by reverse transfection. J Nanosd Nanotechnol 10 3170-3174... 

In this chapter we describe protocols for reverse transfection to generate mammalian cell arrays for systematic gene knock-downs by RNAi or knock-ins by ectopic cDNA expression. The method is suitable for high content screening microscopy at a high spatial and temporal resolution allowing even time-lapse analysis of hundreds of samples in parallel. 

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