A microneedle vaccine printer for thermostable COVID-19 mRNA vaccines
. Lipids were dissolved in ethanol at a molar ratio of 35:16:46.5:2.5 cKK-E12:DOPE:cholesterol:C14-PEG2000 or 38.4:12.3:47.4:1.9 Lipid 5:DOPE:cholestorol:C14-PEG2000 when using cKK-E12 or Lipid 5 as the ionizable lipid, respectively. To prepare the LNPs, the ethanoic solution was rapidly added to and mixed with an mRNA solution buffered with citrate at pH 3 at volume ratio 3:1 . When using cKKe12, the ionizable lipid-to-mRNA weight ratio was set to 10, and the final mRNA concentration was 0.
For the in vitro evaluation of MNPs loaded with LNPs, four MNPs were fabricated using the two-step loading, 10 µg of mRNA and 8 mg PVP:PVA for the tip loading. Two MNPs were used to quantify LNP tip loading, and two MNPs were used to transfect HeLa cells. Needles were cut and dissolved in 1 ml of TE buffer for the RNA quantification and in 1 ml of DMEM for the cell transfection.
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