CyTOF analysis was performed by PLTTech Inc. (Hangzhou, China) according to the protocol described previously.
33 (link) In brief, mouse liver tissue was dissociated into single cells with DNAase, collagenase IV, and hyaluronidase (Sigma‐Aldrich, Saint Louis, MO, USA). Immune cells were concentrated using Percoll density gradient media (Sigma‐Aldrich), while red blood cells were removed using ACK Lysing Buffer (Sigma‐Aldrich). Qualified samples were gathered in blocks and stained for 30 min with a panel of 42 antibodies, followed by overnight fixation. Permeabilisation buffer was applied, and the cells were incubated in an intracellular antibody mixture. The cells were rinsed, and the signals were detected using a CyTOF system (Helios, Fluidigm, South San Francisco, CA, USA). The types of immune cells were identified via nonlinear dimensionality reduction (viSNE) followed by density‐based clustering.
33 (link) In brief, mouse liver tissue was dissociated into single cells with DNAase, collagenase IV, and hyaluronidase (Sigma‐Aldrich, Saint Louis, MO, USA). Immune cells were concentrated using Percoll density gradient media (Sigma‐Aldrich), while red blood cells were removed using ACK Lysing Buffer (Sigma‐Aldrich). Qualified samples were gathered in blocks and stained for 30 min with a panel of 42 antibodies, followed by overnight fixation. Permeabilisation buffer was applied, and the cells were incubated in an intracellular antibody mixture. The cells were rinsed, and the signals were detected using a CyTOF system (Helios, Fluidigm, South San Francisco, CA, USA). The types of immune cells were identified via nonlinear dimensionality reduction (viSNE) followed by density‐based clustering.
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