Mouse ES cells (E14TG2a) were maintained on gelatin-coated dishes in ATCC-formulated Dulbecco’s Modified Eagle’s Medium (ATCC), supplemented with 0.1 mM of 2-mercaptoethanol, 10% fetal bovine serum (StemCell Technologies), 2 mM L-glutamine, 0.1 mM MEM nonessential amino acid, 100 U/mL penicillin, 10 µg/mlstreptomycin, and 10 ng/mL LIF (StemCell Technologies). The mES cells were passaged every 2 d at a ratio of 1:5 by washing with PBS, dissociating with 0.25% trypsin (GIBCO) for 3 min at 37°C, and resuspending in mES media. Media was changed daily. To induce differentiation, the mES cells were passaged and then cultured in ES cell culturing media without LIF. Media was changed every 2 d. The undifferentiated (E14-d0) and differentiating (E14-d6) states of mES cells were verified by SSEA-1 (stage-specific embryonic antigen-1) staining with StainAlive SSEA-1 Antibody (DyLight 488) (Stemgent) and quantitative RT-PCR analysis of relative expression levels of three major pluripotency factors including Nanog, Sox2, and Pou5f1.
Mem nonessential amino acid
Manufactured by STEMCELL
MEM nonessential amino acid is a laboratory reagent used in cell culture media formulations. It provides a source of nonessential amino acids to support cell growth and proliferation in vitro.
Automatically generated - may contain errors
Lab products found in correlation
L glutamine, by STEMCELL (2 mentions)
Fetal bovine serum (fbs), by STEMCELL (1 mentions)
Penicillin, by STEMCELL (1 mentions)
Trypsin, by Thermo Fisher Scientific (1 mentions)
Sodium pyruvate, by Beyotime (1 mentions)
Interleukin 2 (il 2), by Thermo Fisher Scientific (1 mentions)
Hepes, by STEMCELL (1 mentions)
Anti cd28 antibody, by BioLegend (1 mentions)
Cd8 t cell isolation kit, by BioLegend (1 mentions)
Anti cd3 antibody, by BioLegend (1 mentions)
2 protocols using mem nonessential amino acid
1
Maintaining and Differentiating Mouse Embryonic Stem Cells
2
Isolation and Activation of Murine CD8 T Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Isolation of CD8 T cells from the spleens of wild-type C57BL/6 mice was carried out using the CD8 T Cell Isolation Kit (BioLegend, #480035) in accordance with the manufacturer’s instructions. For preactivation, purified CD8 T cells were quantified and seeded onto a flat-bottom 96-well culture plate precoated with 2 µg/mL of anti-CD3 antibody (BioLegend, #100340) and 2 µg/mL of anti-CD28 antibody (BioLegend, #102116). The cells were cultured in a comprehensive medium comprising RPMI 1640, supplemented with 10% FBS, 100 U/mL interleukin-2 (Peprotech, #212–12), 2 mM L-glutamine (STEMCELL Technologies, #07100), 50 µM β-mercaptoethanol (GENOM Biotech, #GNM21985-1), 1 mM sodium pyruvate (Beyotime, C0331), 100 µM MEM non-essential amino acid (STEMCELL Technologies, #07600), and 10 mM HEPES (STEMCELL Technologies, #07200). After 24 hours, the CD8 T cells were co-cultured with macrophages at a 1:1 ratio. Following an additional 48-hour co-culture period, CD8 T cells were subjected to staining for interferon-γ (IFN-γ) and granzyme B (GZMB) to assess their activation status.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!