Mice were anesthetized with sodium pentobarbital (50 mg/kg), and after collecting a blood sample, the liver was harvested and placed in a 15-mL tube containing 3 mL of tissue storage medium (cat# 130–100-008, Miltenyi Biotec, Bergisch Gladbach, Germany) on ice. The dissociation was performed with the Multi Tissue Dissociation Kit (#130–110-201, Miltenyi Biotec) following the manufacturer’s specifications, using the right liver lobule and 5 mL of working volume. Once red blood cells were removed, the cell pellet was resuspended in 500 µl of RPMI before adding 500 µl of cisplatin/rhodium intercalator solution [10 µM of cisplatin (natural abundance), 4 µM of rhodium intercalator (cat# 201103A, Fluidigm, San Francisco, CA, USA) in RPMI 1640] for live/dead discrimination for 5 min RT before being quenched by 5 mL of RPMI 1640 (10% FBS, 1% DNase). Finally, cells were fixed in RPMI 1640 with 1.6% PFA for 10 min and transferred to cryotubes. The pellets were stored at −80°C.
Rhodium intercalator
Manufactured by Standard BioTools
Sourced in United States
The Rhodium-intercalator is a laboratory instrument designed to detect and analyze nucleic acid sequences. It functions by intercalating with DNA or RNA, which allows for the identification and quantification of specific genetic material. The Rhodium-intercalator provides a reliable and precise method for researchers to study genomic and transcriptomic targets.
Automatically generated - may contain errors
Lab products found in correlation
Cytof2 mass cytometer, by Standard BioTools (2 mentions)
Cell strainer, by BD (2 mentions)
Eq four element calibration beads, by Standard BioTools (2 mentions)
0.1 μm spin column, by Merck Group (2 mentions)
Multi tissue dissociation kit, by Miltenyi Biotec (1 mentions)
Saponin, by Merck Group (1 mentions)
Iridium intercalator, by Standard BioTools (1 mentions)
4 protocols using rhodium intercalator
1
Liver Dissociation and Cell Preparation
2
Mass Cytometry for Multiplexed Cellular Analysis
3
Mass Cytometry for Multiplexed Cellular Analysis
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To eliminate technical variability during staining or acquisition, individual samples within one experiment were palladium-barcoded as described previously63 and combined into a composite sample before further processing and staining. Cell-surface antibody master-mix in CSM was filtered through a pre-wetted 0.1 μm spin-column (Millipore) to remove antibody aggregates and added to the samples. After incubation for 30 min at RT, cells were washed once with CSM. To enable intracellular staining, cells were permeabilized by incubating with ice-cold MeOH for 10 min on ice and washed to times with CSM to remove any residual MeOH. Intracellular antibody master-mix in CSM was added to the samples and incubated for 1 h at RT. Cells were washed once with CSM and resuspended in intercalation solution (1.6% PFA in PBS and 0.5 μM rhodium-intercalator (Fluidigm)) for 20 min at RT or overnight at 4 °C. Before acquisition, samples were washed once in CSM and twice in ddH2O and filtered through a cell strainer (Falcon). Cells were then resuspended at 1 × 106 cells/mL in ddH2O supplemented with 1x EQ four element calibration beads (Fluidigm) and acquired on a CyTOF2 mass cytometer (Fluidigm).
4
Cellular Surface Staining for CyTOF
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All surface staining was performed in CSM for 30 min at RT. Where indicated, cells were fixed with 1.6% PFA in PBS for 10 min at RT and permeabilized with MeOH for 10 min on ice. Before acquisition, samples were resuspended in intercalation solution (1.6% PFA in PBS, 0.02% saponin (Sigma) and 0.5 μM iridium-intercalator or 0.5 μM rhodium-intercalator (both Fluidigm)) for 1 h at RT or overnight at 4 °C.
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