Wildtype (WT) MEFs were prepared as described previously.21 PECs were prepared 4 days after injection of 2 mL of 3% thioglycollate broth (Becton Dickinson, Franklin Lakes, NJ, USA) by peritoneal lavage with ice‐cold phosphate‐buffered saline (PBS). Cells were washed twice and used for total RNA preparation or immunoblotting. WT MEFs, PECs, HeLa cells, Tig3 cells, and RAW264 cells were cultured in Dulbecco’s modified Eagle’s medium (Nissui Pharmaceutical Co., Tokyo, Japan) supplemented with 10% fetal bovine serum and 50 µg/mL kanamycin (Meiji Seika Pharma, Tokyo, Japan). Human acute monocytic leukemia THP‐1 cells (Health Science Research Resources Bank, Osaka, Japan) were cultured in RPMI medium (Nissui Pharmaceutical Co.) supplemented with 5% fetal bovine serum.
Rpmi medium
Manufactured by Nissui Pharmaceutical
Sourced in Japan
RPMI medium is a cell culture medium commonly used for the growth and maintenance of various mammalian cell lines. It provides the necessary nutrients, salts, and buffers to support the in vitro cultivation of cells. The core function of RPMI medium is to create a controlled and optimal environment for cell growth and proliferation.
Automatically generated - may contain errors
Lab products found in correlation
Thioglycollate broth, by BD (1 mentions)
Dulbecco s modified eagle s medium, by Nissui Pharmaceutical (1 mentions)
Kanamycin, by Meiji Seika Pharma (1 mentions)
Lps from escherichia coli o111 b4, by Merck Group (1 mentions)
Thioglycollate medium, by BD (1 mentions)
12 well plates, by Iwaki (1 mentions)
2 protocols using rpmi medium
1
Isolation and Culture of Mouse Cells
2
Peritoneal Macrophage Isolation and Stimulation
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Thioglycollate medium (Becton Dickinson, Sparks, MD) (1 mL/20 g of body weight) was intraperitoneally injected into mice, and peritoneal exudate cells and fluids were collected on day 2 and 4 by washing the cavity with 8 mL of PBS as described previously9 (link). Cell number was determined by Trypan Blue exclusion. Cytocentrifuge preparations were Giemsa-stained, and cell subsets were identified and counted. For preparation of MΦs, the peritoneal cells were washed, counted and then seeded into 12-well plates (Iwaki Glass, Tokyo, Japan) at a cell density of 106 cells/mL in 1 mL of RPMI medium (Nissui, Tokyo, Japan) supplemented with 10% (v/v) fetal calf serum. After incubation for 2 hours in a CO2 incubator, the supernatants and non-adherent cells were removed. More than 90% of adherent cells were peritoneal MΦs. The cells were then incubated with or without 10 μg/mL LPS from Escherichia coli O111:B4 (Sigma, St. Louis, MO) in medium containing 2% serum for 24 hours. The culture media were taken for measurements of prostanoids.
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