Nucleated cells from the femur and tibia of 3-week post-pIpC mice were lineage depleted with a MACS lineage cell separation kit according to manufacturer’s instructions (Miltenyi Biotec, Auburn, CA). Lineage depleted cells were cultured onto 30,000 OP9 cells (plated night before) in IMDM supplemented with 10% defined FBS, 55mM BME, 50 U/ml penicillin, 50mg/mL streptomycin, 0.1mM Glutamax, 10ng/mL Flt3L, and 5ng/mL IL-7. Recombinant mouse cytokines were obtained from R&D Systems (Minneapolis, MN) or Invitrogen (Carlsbad, CA). Cells were transferred onto fresh OP9 cells every 3 days. To evaluate myeloid and B cell differentiation, cells were analyzed 6 days after culture with B220-APC and CD11b-APC/cy7 antibodies (BioLegend, San Diego, CA).
Macs lineage cell separation kit
Manufactured by Miltenyi Biotec
The MACS lineage cell separation kit is a laboratory equipment designed for the isolation and purification of specific cell types from complex biological samples. It utilizes magnetic-activated cell sorting (MACS) technology to effectively separate target cells based on their unique surface markers or intracellular characteristics. The core function of this kit is to enable the isolation of desired cell populations for further analysis or downstream applications.
Automatically generated - may contain errors
4 protocols using macs lineage cell separation kit
1
Lineage Depletion and Cell Culture Protocol
2
Myeloid and B-cell Differentiation Assay
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Nucleated cells from the femur and tibia of 5–6 week old mice were lineage depleted with a MACS lineage cell separation kit according to manufacturer’s instructions (Miltenyi Biotec, Auburn, CA). Lineage depleted cells were cultured onto 30,000 OP9 cells (plated night before) in IMDM supplemented with 10% defined FBS, 55mM BME, 50 U/ml penicillin, 50mg/mL streptomycin, 0.1mM Glutamax, 5ng/mL Flt3L, and 1 ng/mL IL-7. Cultures were treated with DMSO control, 1 uM of BMP inhibitor LDN 193189 (Abcam, Cambridge, MA), or 100nM FoxO1 inhibitor AS1842856 (Calbiochem, Billerica, MA). Mouse cytokines were obtained from R&D Systems or Invitrogen. Cells were transferred onto fresh OP9 cells every 3 days. To evaluate myeloid and B-cell differentiation in cultures, cells were analyzed 6 or 9 days after culture with B220-APC and CD11b-APC/cy7 antibodies (BioLegend, San Diego, CA) by flow cytometry on the Beckman Coulter FC500 flow cytometer.
3
Bone Marrow Cell Differentiation Assay
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The use of mice in these experiments was approved by the University of New Mexico LACUC (Protocol #07UNM027). In vitro culture of lineage negative bone marrow cells was performed as previously described [30 (link)]. Lineage negative bone marrow cells prepared with MACS lineage cell separation kit (Miltenyi Biotec). Cells were infected with MigR1 or MigR1-E47 retroviral supernatant. After infection cells were cocultured with OP9 cells in IMDM media containing 1 ng/mL IL-7 and 5 ng/mL Flt3L. After 12 d of coculture differentiation cells were stained with CD19-PE and CD11b-APC (Invitrogen) and analyzed with a FacsCalibur flow cytometer (Becton Dickinson).
4
Hematopoietic Lineage Commitment Assay
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Nucleated cells from the femur and tibia of 5-to 6-wk-old mice were lineage depleted with a MACS Lineage Cell Separation Kit, according to the manufacturer's instructions (Miltenyi Biotec). Lin 2 cells were cultured onto 40,000 OP9 cells (plated the night before) in IMDM, supplemented with 10% defined FBS, 55 mM 2-ME, 50 U/ml penicillin, 50 mg/ml streptomycin, 0.1 mM GlutaMAX, 5 ng/ml Flt3L, and 1 ng/ml IL-7. EML cells overexpressing MSCV, or MSCV-mirn23a were cultured onto 40,000 OP9 cells (plated the night before) in IMDM, supplemented with 20% FBS, 10% COS-KSL conditioned media (source of SCF), 5 ng/ml Flt3L, and 1 ng/ml IL-7. Recombinant mouse cytokines were obtained from R&D Systems or Thermo Fisher Scientific Life Sciences. Cells were transferred onto fresh OP9 cells every 3 d. To evaluate myeloid and B cell differentiation in nontransduced cultures, cells were analyzed 6 d after culture with B220-APC and CD11b-APC/ Cy7 antibodies (BioLegend). In MSCV or MSCV-Trib3-overexpressing hematopoietic cultures, cells were analyzed 9 d after OP9 culture.
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!