PBMCs and splenocytes obtained by Ficoll-Hypaque density centrifugation were stained with the following anti-human antibodies: V500 anti-CD19 (Becton Dickinson), PerCPcy5.5 anti-CD38 (Biolegend), PE-cy7 anti-CD24 (Biolegend), FITC anti-IgD (Invitrogen), APC anti-CD27 (Becton Dickinson). Dead cells were excluded by Sytox Blue staining (Invitrogen). B cells (1,000-20,000) were sorted using a FACSAria (Becton Dickinson) and collected into 30 μl of RT lysis buffer(14 (link)).
Pe cy7 anti cd24
Manufactured by BioLegend
The PE/Cy7-anti-CD24 is a fluorescent-conjugated antibody that specifically binds to the CD24 cell surface antigen. It can be used in flow cytometry applications to identify and quantify CD24-expressing cells.
Automatically generated - may contain errors
Lab products found in correlation
Anti cd44 apc, by BD (2 mentions)
Hank s balanced salt solution, by Thermo Fisher Scientific (2 mentions)
Bovine serum albumin (bsa), by Merck Group (2 mentions)
Facsvantage se, by BD (2 mentions)
Sytox blue, by Thermo Fisher Scientific (1 mentions)
Facsaria, by BD (1 mentions)
Anti epcam apc, by BioLegend (1 mentions)
4 protocols using pe cy7 anti cd24
1
Isolating and Sorting B Cells from PBMCs
2
Single-cell Isolation and Staining
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To obtain single cell suspension, rather than clumps, the tissue was incubated for 10 min on ice in a 10 mM EDTA solution, before to be cut in small pieces and moved for other 10 min in a pre-warmed 10 mM EDTA solution. The tissue was shaked vigorously every 2 min. Cells were filtered through a 70 μm cell strainer and spun down at 300 × g for 5 min at 4 °C. Cells were resuspended in FACS buffer and stained with combination of APC-anti-Epcam (1:100, BioLegend, 118214) and PE/Cy7-anti-CD45 (1:1000, Biolengend, 103114) or APC-anti-Epcam and PE/Cy7-anti-CD24 (1:100, BioLegend, 101821). FcX blocking solution (BioLegend) was added at a dilution of 1:50.
3
Cytometric Isolation of CD44 Subpopulations
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Flow cytometry and FACS were performed as described previously 37 (link). To determine CD44high-CD24low/- cells (CD44H) and CD44low/--CD24low/- cells (CD44L) subpopulations, cells were suspended in Hank's balanced salt solution (Life Technologies) containing 1% BSA (Sigma-Aldrich) and stained with PE/Cy7-anti-CD24 at 1:10 (BioLegend, San Diego, CA) and APC-anti-CD44 at 1:20 (BD Biosciences) on ice for 30 min. FACS Vantage SE (BD Biosciences) was used to isolate CD44L and CD44H cells. Flow cytometry was repeated for each genotype and condition at least three times.
4
Cytometric Isolation of CD44 Subpopulations
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