Leveraging mRNAs sequences to express SARS-CoV-2 antigens in vivo

Chunxi Zeng; Xucheng Hou; Jingyue Yan; Chengxiang Zhang; Wenqing Li; Weiyu Zhao; Shi Du; Yizhou Dong

Immunostaining and microscopy imaging 293T cells were grown overnight on cover slides placed in a 6-well plate. TT3 nanoparticles were formulated with mRNAs encoding SARS-CoV-2 antigens, S, RBD, M, N, and E proteins, as described above. Six hours after treatment, cells were washed with phosphate buffered saline (PBS) and fixed with 10% formalin, followed by permeabilization with 0.2% Tween-20. Staining was conducted using primary rabbit anti-FLAG antibody (abcam, ab1162) at 1:200 dilution and FITC-linked secondary goat antirabbit polyclonal antibody (abcam, ab6717) at 1:1000 dilution. Nucleus was stained with Hoechst 33342 (Thermo). After sealing the slides, images were taken on a Nikon Eclipse Ti Inverted Fluorescence Phase Contrast Microscope with NIS-Elements BR imaging software (version 4.20).