Multiple transfection

Prepare transfection mix A that contains 1 fig of Renilla luciferase target plasmid (e.g., p-R-luc-4 sites) and 100 /(I of optiMEM (Invitrogen, CA). This is enough transfection mixture for 1 well, and can be scaled up for multiple transfections. For cultures that are to receive miR duplex, add 2 nM (calculated for a final volume of 500 fiL for each well). 

Utilization of Double (Multiple) Transfected Cells for the Characterization of Hepatobiliary Transport... 

The main advantages of the biolistic transfection are (1) the need of a limited quantity of DNA compared to other transfection methods (2) the possibility to very easily perform multiple transfections (co-transfection) with different DNAs [24] (3) its high efficiency in post-mitotic neurons and (4) its optimal applicability to organotypic cultures. 

Use 96-well conical bottom PCR plates to prepare multiple transfection mix pools. Using a 96-well plate will facilitate ease of pipetting and organization overall. In order to screen a 1,000-compound library, use 72 transfection mix pools (six transfection mix pools per plate) and transfect cells sufficient to plate twelve 10 cm dishes (in other words 72 wells of the PCR plate) see Note 5). 

Therefore, evidence exists to support numerous possible mechanisms for the uptake promoted by TAT peptides. An alternative explanation is that TAT is a sticky opportunistic peptide that has the ability to bind to the cell surface and exploit multiple mechanisms in order to enter the cytoplasm (92). Clearly, the mechanism of internalization requires further study. Nevertheless, it has been observed that transfection with CPPs requires less lipid and therefore proves to be less cytotoxic to cells in vitro and in vivo, making it a promising vector system for future gene therapy (95,104). 

Besides Cx40, Cx43 and Cx45, Cx37 channels are expressed in the cardiovascular tissue. These channels are frequently found in endothelium. These channels were expressed in human Cx37-transfected neuroblastoma cells (N2A cells) and exhibited a pronounced voltage dependence and multiple conductance states [Reed et al., 1993]. Several single-channel conductances were found ... 

Garcia, M., Sakamoto, K., Shigekawa, M., Nakanishi, S., Ito, S. Multiple mechanisms of arachidonic acid release in Chinese hamster ovary cells transfected with cDNA of substance P receptor, Biochem. Pharmacol. 1994, 48, 1735-1741. 

One of the most important criteria of transfection with polycationic polymers is the ability of the polymer to condense plasmid DNA. Naked DNA can be taken up by cells and expressed, however not at sufficiently high levels, presumably due to degradation at one or multiple points in the transfection process. In solution,... 

Table 21.4 Level of gene expression by co-transfection of animals with multiple genes ...

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