BMDMs were isolated as previously described (23 (link)). In short, mice (8~14 weeks old) were sacrificed and both legs were sterilized in 70% ethanol. Attached tissues were removed by dissection with scissors and the isolated bones were flushed or crushed twice in a mortar with 5 mL RPMI 1640 medium. The suspension from crushed bones was filtered through 100 μm cell strainer (Falcon#352360) to remove debris. Filtered cells were centrifuged at 500 x g, 15 mins at room temperature. Supernatant was removed and cell pellets were resuspended in RPMI 1640 (+M-CSF 1, 100 ng/mL). The cell suspension (from 1 leg) was divided into four Nunc Square 25-cm dishes (Thermo Scientific#166508), with each plate containing 10 mL cell suspension. On Day 3 post-seeding, 5 mL fresh RPMI 1640 (+M-CSF 1) medium was added to each plate. On Day 6, BMDMs were ready for collection or re-seeding for new experiments. All mice handling was done according to Swiss federal guidelines for animal experimentation and approved by the FMI Animal committee and the local veterinary authorities (Kantonales Veterinäramt of Kanton Basel-Stadt).
Nunc square 25 cm dishes
Manufactured by Thermo Fisher Scientific
The Nunc Square 25-cm dishes are lab equipment designed for cell culture applications. They provide a square-shaped surface area of 25 square centimeters for growing cells in a controlled laboratory environment.
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Lab products found in correlation
2 protocols using nunc square 25 cm dishes
1
Isolation and Culture of Bone Marrow-Derived Macrophages
2
Isolation of Bone Marrow-Derived Macrophages
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BMDMs were isolated as previously described (26 (link)). In short, mice (8∼14 weeks old) were sacrificed and both legs were sterilized in 70% ethanol. The attached tissues were removed by dissection with scissors and the isolated bones were flushed or crushed twice in a mortar with 5 ml RPMI 1640 medium. The suspension from crushed bones was filtered through 100 μm cell strainer (Falcon#352360) to remove debris. Filtered cells were centrifuged at 500g, 15 min at room temperature. The supernatant was removed and cell pellets were resuspended in RPMI 1640 (+M-CSF 1, 100 ng/ml). The cell suspension (from one leg) was divided into four Nunc Square 25-cm dishes (Thermo Scientific#166508), with each plate containing 10 ml cell suspension. On Day 3 post-seeding, 5 ml fresh RPMI 1640 (+M-CSF 1) medium was added to each plate. On Day 6, BMDMs were ready for collection or re-seeding for new experiments. All mice handling was done according to Swiss federal guidelines for animal experimentation and approved by the FMI Animal Committee and the local veterinary authorities (Kantonales Veterinäramt of Kanton Basel-Stadt).
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