c-Kit+ cells were isolated from mouse bone marrow cells and cSAM cells using MACS (magnetic-activated cell sorting), and were cultured in Methocult 3231 (StemCell Technologies, Vancouver, BC, Canada) supplemented with 20 ng/ml SCF and 10 ng/ml mouse IL-3. A total of 300 cells were plated in duplicates for each round of plating. Colony counting and replating were performed every 5 days.
Methocult 3231
Manufactured by STEMCELL
Sourced in Canada
Methocult 3231 is a methylcellulose-based medium designed for the culture and enumeration of human and murine hematopoietic progenitor cells. It provides a semi-solid matrix that supports the growth and differentiation of these cells.
Automatically generated - may contain errors
4 protocols using methocult 3231
1
Expansion of c-Kit+ Hematopoietic Cells
2
Hematopoietic Cell Colony Assays
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
For CFU-Mk and CFU-Myeloid assays, 50 × 103 cells were cultured in methylcellulose media (MethoCult 3231; Stemcell Technologies) in the presence of recombinant mouse (m) SCF (50 ng/ml), mTPO (50 ng/ml), human (h) EPO (5 U/ml), mIL-3 (20 ng/ml) and mIL-11 (40 ng/ml; cytokines from PeproTech), as described previously (Akashi et al., 2000 (link)). For CFU–preB assays, 15 × 104 cells were cultured in methylcellulose media containing hIL-7 (MethoCult 3630; Stem Cell Technologies). All assays were performed in duplicate, and CFUs were counted after 7 d. Single colonies were cytospun onto slides, subjected to May–Grünwald–Giemsa (MGG) staining, and analyzed with an upright microscope (DM4000 B 40X; Leica). Images were captured with the Coolsnap software.
3
Methylcellulose Assay for Murine and THP-1 Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
For methylcellulose medium, 20 ml IMDM (Life Technologies) was added to 80 ml MethoCult 3231 (STEMCELL Technologies, Catalog #03231), vortexed, and allowed to settle. For primary murine cell lines, the methylcellulose was supplemented with cytokines 20 ng/ml SCF, 10 ng/ml IL-3, 10 ng/ml IL-6 and 10 ng/ml GM-CSF. No antibiotics were added. Cells (THP-1 cells, MLL-AF9 resultant drug compound concentration 0.1%). The plate was vortexed, and the side troughs and unused wells were half filled with PBS (Sigma) to prevent edge effects due to uneven evaporation.
Plates were incubated at 37°C 5% CO 2 (day 0), and then scanned on day 6 (murine cells), or day 9 (THP-1 cells).
Plates were incubated at 37°C 5% CO 2 (day 0), and then scanned on day 6 (murine cells), or day 9 (THP-1 cells).
4
Hematopoietic Progenitor Cell Assay
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Whole BM cells or cells sorted on the basis of surface markers were plated in methylcellulose (Methocult 3231; Stem Cell Technologies), supplemented with 20 % IMDM, 1 % P/S, 50 ng/ml mSCF, 10 ng/ml mIL-3 and 10 ng/ml hIL-6, with or without
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!