Natural Killer Cell Isolation from Liver Tissue for Selective Expansion
Transcrição
To isolate natural killer cells, or NK cells — specialized white blood cells involved in innate immunity, begin with human liver tissue containing diverse cell populations, including fat-containing cells, erythrocytes, granulocytes, lymphocytes, and other associated cells entrapped within a collagen-rich extracellular matrix or ECM.
Mince the tissue into small pieces and transfer them to a tube containing collagenase — a proteolytic enzyme. Collagenase cleaves the collagen, disrupts the matrix, and loosens the ECM-bound cells.
Insert the tube into a tissue dissociator. This mechanical dissociation further disintegrates ECM to release loosely attached cells into the solution. Filter the contents to remove undigested tissue and ECM fragments and collect the filtrate containing various cells.
Resuspend the cells into a buffer containing polyvinylpyrrolidone or PVP-coated silica beads. The PVP beads selectively bind with fat-containing cells, effectively separating them from the rest of the cells.
Centrifuge at low speed to pellet the cell population; discard the supernatant containing debris and bead-bound fat cells. Resuspend the pellet and overlay the cell suspension onto a tube containing a density gradient medium to separate the cells based on their densities.
Centrifuge at low speed. The higher-density erythrocytes and granulocytes settle at the bottom, while lymphocytes appear at the interphase between two liquids.
Collect the cells from interphase and culture them in a selective medium for NK cell expansion.
