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Do siRNAs synthesized using 2'-ACE chemistry need to be HPLC purified for in vitro use? For in vivo use?
2'-ACE synthesis chemistry has very high coupling efficiencies resulting in siRNAs of high quality and purity. If purification is needed, such as with siRNAs modified with dyes or bioconjugates, we use an optimized PAGE method to improve overall yield and provide duplexes with ~95% purity. In general, the majority of siRNAs ordered for use in in vitro assays do not require purification.
siRNAs processed for in vivo use include an HPLC purification option. In this instance, HPLC is the method of choice in order to maintain the correct counter ion (Na+). Although many in vivo studies have not used purified duplexes, different experimental systems may require very pure preparations of siRNAs. We encourage investigators to consult PubMed (http://www.ncbi.nlm.nih.gov/entrez/query.fcgi) or HighWire (http://highwire.stanford.edu) for similar studies to determine specific purity needs.