LSK immunophenotype hematopoietic stem cells after irradiation at day 2

To investigate the impact of radiation damage on hematopoietic stem cell fate decision, we conducted a longitudinal study, collecting single-cell data on days 2 post-radiation exposure. This approach enabled us to observe changes in the subpopulations of hematopoietic stem cells at the LSK level in mice over time. Additionally, we assessed alterations in their fate determination and lineage differentiation tendencies throughout the recovery process. To perform 10X Genomics single-cell sequencing on mouse bone marrow LSK cells, begin by euthanizing the mouse following ethical guidelines and extracting the femurs and tibias. Flush the bone marrow using a syringe with buffer PBS with 2% FBS, to obtain a single-cell suspension. Filter this suspension through a 40 µm cell strainer to remove debris and clumps. Next, enrich the LSK (Lineage^-, Sca-1^+, c-Kit^+) cells using fluorescence-activated cell sorting (FACS). Count the cells and assess viability using trypan blue staining. Adjust the concentration of LSK cells to the required density 700-1200 cells/µL, for optimal performance in the 10X Genomics Chromium Controller. Load the single-cell suspension, reagents, and gel beads into the appropriate wells of a 10X Genomics Single Cell 3' Chip and run the chip in the Chromium Controller to encapsulate single cells into nanoliter-scale Gel Bead-In-EMulsions (GEMs). Perform reverse transcription within the GEMs to generate barcoded cDNA from the mRNA of individual