Endotoxin is a potent stimulator of the mammalian immune system in vivo, and it may decrease tissue culture cell viability and inhibit transfection efficiency in vitro. Significant amounts of endotoxin are released from E. coli during plasmid purification. For maximum consistency and when transfecting a broad variety of cell lines, it is best to use plasmid DNA containing as little endotoxin as possible. The system includes a proprietary Endotoxin Removal Resin, based on a novel magnetic capture technology, to purify plasmid DNA to low levels of endotoxin contamination. For supposedly purified plasmid DNA, methods have been described to remove endotoxin from these samples.
Precipitation removes salt, guanidine and endotoxin from DNA in solution.
Although not a quantitative removal, this method may be sufficient to improve transfection, especially for cells not extremely sensitive to endotoxin. More stringent endotoxin removal from purified DNA uses either extraction or chromatography. At temperatures below 20 C, Triton X-114 detergent will dissolve in aqueous solutions. At temperatures above 20 C, the detergent separates into two phases where lipophilic endotoxin partitions to the organic phase. The eluate is collected and DNA is precipitated with salt and alcohol.