title: GSM8403167: S7: 108 Day, Control, Female; Homo sapiens; RNA-Seq
library_construction_protocol: From each lung, 400 mg of tissue was weighed out and placed on a clean glass petri dish containing chilled HBSS and minced (1-2 mm sized pieces) using a sterile razor blade. Lung was then further dissociated with the Miltenyi gentleMACs dissociator (Cat: 130-093-235) using the manufacturer-set dissociator program. Cell solution was then passed through prepared 70um cell strainer. C tube was then rinsed with 3mL HBSS and passed through strainer, and strainer was rinsed further with an additional 2mL HBSS. Strained cell suspension was then centrifuged at 500G for 5 minutes followed by decanting. Cell pellet was then incubated in Red Blood Cell Lysis Buffer (3 minutes at room temperature), diluted with HBSS (4 HBSS:1 RBC), and centrifuged at 500G for 5 minutes followed by decanting. Cell pellet was subsequently resuspended in 2mL 1% BSA and centrifuged at 500G for 5 minutes followed by decanting (this step was repeated a second time). Cells were then resuspended in 1%FBS DMEM/F12 at a concentration of 1000 cells/μL for single cell RNA library preparation with all subsequent cells frozen down in 90% FBS/10% DMSO solution. Single cell capture and library production was performed on the Chromium 10X Genomics system with version 3 chemistry according to the manufacturer's instructions. Sequencing was performed on an Illumina NovaSeq, with read alignment to GRCh38.