40–80 × 106 frozen spleen cells were thawed into DMEM with 10% fetal calf serum and 100 U/mL penicillin plus 100 µg/ mL streptomycin. Cells were centrifuged and resuspended 50 µL of ice-cold fluorescence-activated cell sorting (FACS) buffer composed of 1× DPBS and 1% newborn calf serum (Thermo Fisher, cat#26010074). PostF PE/DyLight650 conjugated tetramers were added at a final concentration of 25 nM in the presence of 2% rat and mouse serum (Thermo Fisher) and incubated at room temperature for 10 min. S2P PE tetramers were then added at a final concentration of 5 nM and incubated on ice for 25 min, followed by a 10 mL wash with ice-cold FACS buffer. Next, 50 μL of anti-PE-conjugated microbeads (Miltenyi Biotec, Auburn, CA, USA, cat#130-090-855) were added and incubated on ice for 30 min, after which 3 mL of FACS buffer was added and the mixture was passed over a magnetized LS column (Miltenyi Biotec, cat#130-042-401). The column was washed once with 5 mL ice-cold FACS buffer and then removed from the magnetic field and 5 mL ice and 5 mL ice-cold FACS buffer was forced through the unmagnetized column twice using a plunger to expel cells retained in the column as the bound cell fraction.
Anti pe conjugated microbeads
Manufactured by Miltenyi Biotec
Sourced in United States
Anti-PE-conjugated microbeads are magnetic beads that are conjugated with antibodies specific to the phycoerythrin (PE) fluorescent label. These microbeads can be used to magnetically separate and enrich cells or other target molecules that are labeled with PE.
Automatically generated - may contain errors
Lab products found in correlation
Neural tissue dissociation kit, by Miltenyi Biotec (2 mentions)
Pe conjugated anti cd11b antibody, by Miltenyi Biotec (2 mentions)
Ms column, by Miltenyi Biotec (2 mentions)
Anti cd11b pe, by Miltenyi Biotec (2 mentions)
Percoll, by Santa Cruz Biotechnology (2 mentions)
Anti pe microbeads, by Miltenyi Biotec (2 mentions)
Newborn calf serum, by Thermo Fisher Scientific (1 mentions)
Rat and mouse serum, by Thermo Fisher Scientific (1 mentions)
Dylight 650, by Thermo Fisher Scientific (1 mentions)
Ficoll paque gradient, by GE Healthcare (1 mentions)
Easysep isolation kits, by STEMCELL (1 mentions)
Human b cll cell isolation kit, by Miltenyi Biotec (1 mentions)
4 protocols using anti pe conjugated microbeads
1
Enrichment of Antigen-Specific T Cells
2
Isolating CD11b+ Cells from Mouse Brain Tissues
3
Isolating CD11b+ Cells from Mouse Brain Tissues
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Mouse brain tissues or GBM xenografts were digested with the Neural Tissue Dissociation Kit (Miltenyi Biotec) followed by MACS sorting with PE conjugated anti-CD11b antibody (Miltenyi Biotec, 130-091-240, 1:11) and anti-PE conjugated microbeads (Miltenyi Biotec). Briefly, mouse brains or tumors were washed with PBS, minced, and incubated with enzyme mixes. The dissociated brain tissue was passed through a 40μm cell strainer. Tissue was then re-suspended in a 30% Percoll (Santa Cruz) PBS solution and centrifuged at 700g for 10 minutes to remove myelin. Cells were re-suspended in 90μl IMAG buffer (PBS w/o Ca2+/Mg2+, supplemented with 0.5% BSA and 2mM EDTA, pH 7.2) plus 10μl anti-CD11b-PE (Miltenyi) and incubated on ice for 30 minutes. Cells were then washed three times with 1ml IMAG buffer and incubated with 20μl anti-PE microbeads (Miltenyi) plus 80μl IMAG buffer on ice for 15 minutes. After labeling, cells were washed, re-suspended in 500μl IMAG buffer, and applied to MS column (Miltenyi). After wash for 3 times, CD11b+ cells were flushed out of the column by plunger. CD11b+ cells were then cultured in RPMI/1640 medium supplemented with 10% FBS for 1 hour and used for co-injection or lysed for RNA extraction.
4
Isolation and Purification of Immune Cell Populations
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The peripheral blood mononuclear cells (PBMCs) and BM mononuclear cells were isolated using Ficoll-Paque gradients (GE Healthcare). CD8+ or CD3+ T cells were isolated by negative selection using the Easysep isolation kits (Stem Cell Technologies) with a purity of >90% (online supplemental figure 1a, b ). For effector T cell (CD8+CCR7-) isolation, CD8+ T cells were stained with phycoerythrin (PE)-conjugated anti-CCR7 antibody, followed by anti-PE-conjugated microbeads (Miltenyi) with a purity of >90% (online supplemental figure 1c ). B-CLLs were isolated using the human B-CLL Cell Isolation Kit (Miltenyi) with a purity of >90% (online supplemental figure 1d ).
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!