On day 23 after tumor implantation, tumor tissues were resected, chopped into small pieces, and digested in a 37 °C and 5% CO2 incubator for 20 min with fresh RPMI-1640 (Biowest, L0498) containing collagenase type IV (Worthington Biochemical, LS004189, 1 mg/mL), hyaluronidase (Sigma, H6254, 1 mg/mL), and DNase I (Sigma, DN25, 1.5 mg/mL). Digested tissues were minced, filtered through a 70-μm cell strainer, and used to prepare single-cell suspensions according to the manufacturer’s instructions.
Ls004189
Manufactured by Merck Group
LS004189 is a laboratory instrument used for performing various analytical and experimental tasks. It is designed to meet the needs of researchers, scientists, and technicians working in a wide range of scientific disciplines. The core function of this product is to provide reliable and accurate measurements, data collection, and sample processing capabilities to support the research and development activities of Merck Group customers.
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3 protocols using ls004189
1
Tumor Tissue Dissociation Protocol
2
Isolation of Cardiac Fibroblasts from Murine Hearts
3
Isolation of Renal Leukocytes from Mice
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Mice were euthanized under isoflurane anesthesia. Whole kidneys were harvested and stored in cold PBS for less than 30 min until further processing. The tissues were then dissociated into small pieces using razor blades and incubated with digestion solution (collagenase IV (1 mg/ml; Worthington, LS004189) and DNase I (50 µg/ml; Sigma, DN25)) in RPMI 1640 media (Welgene, L0948-500) supplemented with 10% FBS (Biowest, S1480) for 30 min at 37 °C. Single cells were obtained by filtering the solution through 40 μm strainers. Renal leukocytes were further enriched by density-gradient separation. Total kidney cells were resuspended in a 40% Percoll solution (GE Healthcare, 17-0891-01) and layered on top of an 80% Percoll solution. After centrifugation at 2000 rpm for 30 min, cells were collected from the layer between the 40 and 80% Percoll solutions. Red blood cells were removed using RBC lysis buffer (Biolegend, 420302), and the cells were washed with FACS buffer (2% FBS in PBS) for use in further experiments.
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