Mouse large intestine was split longitudinally and trimmed into small pieces. After shocked in cold PBS vigorously, tissues were transferred into cold PBS with 1 mM DTT and incubated in 37°C shaker (10 min at 250 rpm). Then, tissues were transferred into cold PBS with 0.5 M EDTA and 1 M Hepes and incubated in 37°C shaker for two times (10 min at 250 rpm), which was followed by digestion with collagenase IV (10,000 U/ml; Gibco) in RPMI 1640 medium (Gibco) and incubated in 37°C incubator for 45 min. Last, tissue pieces were then vigorously suspended in culture medium containing 10% fetal bovine serum (ExCell) and 1% penicillin-streptomycin (HyClone) in MEM alpha basic medium (Gibco), and the mixture was passed through 70-μm cell strainer (BD Biosciences). The cell suspension was added into culture medium and cultured in 10-cm dishes.
Mem alpha basic medium
Manufactured by Thermo Fisher Scientific
MEM alpha basic medium is a cell culture medium designed for the maintenance and growth of a variety of cell types. It provides the necessary nutrients and components to support cell survival and proliferation in vitro.
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Lab products found in correlation
3 protocols using mem alpha basic medium
1
Isolation of Mouse Intestinal Cells
2
Culturing Human Endometrial Stromal Cells
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The human endometrial stromal cell line (hESC), provided by the Qilu Hospital Laboratory of Shandong University, was cultured in MEM-alpha basic medium (Gibco) with 10% FBS by incubation in 5% CO2 at 37℃.
3
Primary Trabecular Meshwork Cell Isolation and Characterization
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Primary human trabecular meshwork cells were kindly provided by Professor Wei Zhu (Qingdao University, Qingdao, China) [23 (link)]. This study was approved by the Eye Bank Association of America and the Institutional Review Boards of Beijing Tongren Hospital. Nineteen short tandem repeat (STR) loci plus the gender-determining locus, amelogenin, were amplified using STR Multi-amplification Kit (Basic Cognitive Technology, Beijing, China). The cell strain sample was processed using the ABI 3730xl DNA Analyzer. Data were analyzed using GeneMapper3.2 software (Applied Biosystems, Suzhou, China). Appropriate positive and negative controls were run and confirmed for each sample submitted. The STR analyses are presented in Appendix 1. The cells were cultured in MEM Alpha basic medium (Gibco, Grand Island, NY) supplemented with 20% fetal bovine serum (FBS) and 1% penicillin/streptomycin solution (Gibco). The cells were maintained at 37 °C with 5% CO2.
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