I am trying to synthesize the LNP by Pfizer-BioNTech as a control for my experiments, but every method I try, the LNP does not encapsulate my RNA. The DLS and TEM results are good but when I transfect them into cells, there is no RNA translation that can be detected (and the assay is sensitive). Additionally, when I treat the LNP with Triton-X, the amount of RNA before and after LNP lysis appeared to be similar. My lab/institute/university does not have the NanoAssemblr platform, so I have tried to use the thin-film hydration method as well as a DIY microfluidic system using syringes, Y-connector and Tygon tubing. Could anyone advice on what I am doing wrong or suggest alternative protocols? More information about the recommended volumes and/or ratio of organic-to-aqueous phases are also appreciated. Thank you!
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