All materials were used as received. Sodium hydroxide (NaOH), ammonia solution (28%), sodium carbonate (Na2CO3), calcium chloride dihydrate (CaCl2·2H2O), disodium hydrogen phosphate (Na2HPO4), citric acid, acetic acid, ethylene diamine tetraacetic acid (EDTA), absolute ethanol, glutaraldehyde, crystal violet, Coomassie blue, and phosphate buffer saline (PBS) were purchased from Sinopharm Chemical Reagent (China). Poly(allylamine) hydrochloride (PAH; Mw: 17,500), tetraethyl orthosilicate (TEOS), cetyltrimethylammonium bromide (CTAB; Mw: 364.45), dimethylsulfoxide (DMSO), Tris-buffered saline (TBS), epigallocatechin-3-gallate (EGCG), 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide (EDC), cetylpyridinium chloride, agar, sucrose, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), hexamethyldisilazane (HMDS), paraformaldehyde, dexamethasone, β-phosphoglycerol, Vitamin-C, polyformaldehyde, penicillin/streptomycin, and Triton X-100 were obtained from MilliporeSigma (USA). Fetal bovine serum (FBS) and α-modified essential medium (α-MEM) were supplied by Gibco (Australia) and HyClone (USA), respectively.
α modified essential medium α mem
Manufactured by Thermo Fisher Scientific
Sourced in United States
α-modified essential medium (α-MEM) is a cell culture medium used to support the growth and maintenance of a variety of cell types in vitro. It is a modification of the original Eagle's Minimum Essential Medium (MEM), with the addition of the non-essential amino acid, alpha-ketoglutaric acid, which is essential for certain cell types.
Automatically generated - may contain errors
Lab products found in correlation
Fetal bovine serum (fbs), by Thermo Fisher Scientific (4 mentions)
Penicillin, by Thermo Fisher Scientific (4 mentions)
Streptomycin, by Thermo Fisher Scientific (2 mentions)
10 cm culture dishes, by Corning (2 mentions)
L glutamine, by Thermo Fisher Scientific (2 mentions)
Rankl, by Thermo Fisher Scientific (2 mentions)
Ethylenediaminetetraacetic acid, by Sinopharm (1 mentions)
Phosphate buffer saline (pbs), by Sinopharm (1 mentions)
Dimethyl sulfoxide (dmso), by Merck Group (1 mentions)
Paraformaldehyde (pfa), by Merck Group (1 mentions)
Triton x 100, by Merck Group (1 mentions)
Dexamethasone (dex), by Merck Group (1 mentions)
Sucrose, by Merck Group (1 mentions)
Penicillin streptomycin, by Merck Group (1 mentions)
Agar, by Merck Group (1 mentions)
Cetyltrimethylammonium bromide, by Merck Group (1 mentions)
Tris buffered saline (tbs), by Merck Group (1 mentions)
Cetylpyridinium chloride, by Merck Group (1 mentions)
Tetraethyl orthosilicate, by Merck Group (1 mentions)
Epigallocatechin 3 gallate, by Merck Group (1 mentions)
6 protocols using α modified essential medium α mem
1
Synthesis and Characterization of Nanoparticles
2
Expansion of Mouse Bone Marrow Cells
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The mouse BM cells (2 × 107–3 × 107) were flushed from the long bones with 3% fetal bovine serum (FBS) in PBS. A single-cell suspension of all nuclear cells was obtained by passing all BM cells through a 70-μm cell strainer (Bioscience, Dümmer, Germany). Then, 2.5 × 105 cells per square centimeter were seeded into 10-cm culture dishes (Corning, Lowell, MA, USA) and incubated at 37 ℃ in 5% CO2. After 48 h, the cultures were washed with PBS to eliminate the non-adherent cells. The attached cells were cultured for 10–15 d with α-modified essential medium (α-MEM; Gibco-BRL, Gaithersburg, MA, USA) supplemented with 20% FBS (Gibco-BRL), 2 mmol⋅L−1 L-glutamine, 100 U⋅mL−1 penicillin, and 100 mg⋅mL−1 streptomycin (Invitrogen, Gaithersburg, MD, USA). The cell culture protocol and surface marker identification have been described in our previous studies16 (link),58 (link) (Supplementary Table 1 ).
3
Isolation and Culture of Mouse Bone Marrow Cells
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Mouse BM cells (2-3×107) were flushed out from long bones with 3% FBS in PBS. A single-cell suspension of all nuclear cells was obtained by passing all BM cells through a 70 μm cell strainer (Bioscience, Dümmer, Germany). Then, 2.5×105 cells/cm2 were seeded on 10 cm culture dishes (Corning, Lowell, MA, USA) and incubated at 37 ℃ in 5% CO2. After 48 h, the cultures were washed with PBS to eliminate non-adherent cells. The attached cells were cultured for 10 to 15 days with α-modified essential medium (α-MEM; Gibco BRL, Gaithersburg, MA, USA) supplemented with 20% fetal bovine serum (Gibco BRL), 2 mM L-glutamine, 100 U/mL penicillin, and 100 mg/mL streptomycin (Invitrogen, Gaithersburg, MD, USA). The cell culture protocol and surface markers identification are described in our previous studies 2 (link), 6 (link).
4
Osteoclastogenesis Assay Protocol
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Fetal bovine serums (FBS), α-modified essential medium (α-MEM), and penicillin were purchased from Gibco (Gaithersburg, MD, USA). RANKL was purchased from PeproTech (Rocky Hill, NJ, USA), and M-CSF was purchased from R&D Systems (Minneapolis, MN, USA). All primary antibodies as phospho-p38, p38, phospho-ERK, ERK, phospho-JNK, JNK, phospho-Akt, Akt, and c-Fos were obtained from Cell signaling Technology (Danvers, MA, USA). NFATc1 and cathepsin K were obtained from Santa Cruz Biotechnology (Dallas, TX, USA). Trizol and Supercript II Reverse Transcriptase were purchased from Invitrogen (Carlsbad, CA, USA). Calcium phosphate (CaP) coated plates were purchased from Cosmo Bio (Tokyo, Japan). All other chemicals were purchased from Sigma (St. Louis, MO, USA) unless otherwise indicated [24 (link)]. RT-PCR primers are listed in Table 1 .
5
Establishing Cell Lines for Osteoclastogenesis
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Fetal bovine serum (FBS), α-modified essential medium (α-MEM), and penicillin were purchased from Gibco (Gaithersburg, MD, USA). Parental MDA-MB-231 cells were purchased from a Korean Cell Line Bank (South Korea). RAW264.7 macrophage cells were purchased from ATCC (Manassas, VA, USA). RANKL was obtained from PeproTech (Rocky Hill, NJ, USA. pGL4.50 [luc2/CMV/] vector was purchased from Promega (Madison, WI, USA). The antibiotic hygromycin was purchased from Gibco BRL (Gaithersburg, MD, USA). Primary antibodies were purchased from Thermo Fisher Scientific (Waltham, MA, USA), Santa Cruz Biotechnology (Santa Cruz, CA, USA), and Abcam (Cambridge, MA, USA).
6
Cell Culture Conditions for Breast and Bone Cell Lines
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