Spatial transcriptomic analysis of transplanted vascularized lung organoids

To investigate the cell-cell interactions among the transplanted vascularized organoids, we utilized a Slide-seq-based spatial transcriptomics platform to unbiasedly map the spatial distribution of ligand-receptor pairs. Transplanted organoids were freshly collected, immersed in the O.C.T. compound, and immediately frozen on the dry ice. 10X10 Curio Seeker Spatial transcriptomic was performed following the manufacturer’s instruction. This technique was developed based on the Slide-seq technology for high-resolution genome-wide expression analysis at a resolution of 10um diameter in each spot that was densely packed and randomly deposited. Briefly, 5 scrolls of tissue sections of 10um thickness were collected to derive an RNA integrity number (RIN) to test the tissue’s RNA quality. All organoids’ RIN numbers were higher than 8 to pass the quality test. One 10um thickness tissue section was placed on the Curio Seeker tile and mounted with 30um section of CryoCube. Then, the section was processed for hybridization, reverse transcription, and tissue clearance to yield Curio Seeker Bead resuspension. On the second day, 11 cycles of PCR was utilized for second-string synthesis and cDNA amplification. The purified cDNA was applied for Tagmentation with Dual Indexing Primer V2 and consequent library cleanup and QC. The final library was applied for NGS sequencing with NovaSeq6000 following the recommended sequencing depth.