Cell Isolation Monocyte

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Found 12 matching solutions for this experiment

Upstream tips
Briefly, a single-cell suspension containing 1 × 108 PBMCs per mL of cell separation buffer (PBS, 3% FBS, 10mM EDTA) was prepared.
Protocol tips
Cells were then incubated for 10 min with 20 μL of MagniSort™ enrichment antibody cocktail per 100 μL of cell suspension. Cells were washed and resuspended in separation buffer. Then 10 μL of MagniSort™ negative selection beads were added per 100 μL of cell suspension. Following 5 min of incubation, a magnet was used to remove the bead-bound non-monocyte lymphocytes from the monocytes.
Downstream tips
. Enrichment efficiency was determined by flow cytometry staining with anti-human CD14 eFluor 450 (clone 613D, eBioscience) and anti-human CD16 APC (clone CB16, eBioscience).
Upstream tips
Human macrophages were obtained from peripheral blood mononuclear cells (PBMCs)
Upstream tips
Human monocytes were freshly isolated from peripheral blood of ALS patients and HC.
Protocol tips
For negative selection, PBMCs were diluted to 5 × 107 cells/ml and incubated with 50 µl/ml isolation antibody cocktail and 50 µl/mL platelet removal cocktail for 5 min before the addition of 50 µl/ml RapidSpheres. After an additional 5 min of incubation, PBMCs were placed in the magnet for 3 min and the supernatant, containing monocytes, was collected and washed in PBS (1% FCS).
Downstream tips
The monocytes were resuspended in complete maturation medium and placed in the incubator.
Upstream tips
Subjects reported to the laboratory following an overnight fast and had forearm vein blood collected into a 10 ml K2EDTA vacutainer tube.
Downstream tips
Unwanted cells are labelled for removal with bispecific tetrameric antibody complexes recognizing the respective target antigen (CD2, CD3, CD16, CD19, CD20, CD56, CD66b, CD123, or glycophorin A) and dextran-coated magnetic particles.
Downstream tips
Enriched cells were labeled with CD3, CD19, CD20, CD56, CD66b, HLA-DR, CD14, and CD16 antibodies before sorting classical monocytes by FACS AriaII (BD).
Upstream tips
Human neutrophils and monocytes were isolated from venous blood from a healthy female donor. Peripheral blood mononuclear cells were obtained by gradient separation with Ficoll-Paque
Downstream tips
Slan+ monocytes or CD14+ monocytes (2.5 × 104) were thereafter dispensed into U‐bottom 96‐well plates (Corning, Corning, NY, USA), and cultured for 24 h in RPMI 1640 medium containing 10% FBS, in the absence or in the presence of 20% cell‐free supernatants from SW480, SW620, and MCF7 cell lines.
Downstream tips
Macrophages were obtained from human peripheral blood monocytes. To obtain macrophages, monocytes were seeded on culture plastic and incubated for 7 days in DMEM/F12 medium with 2% fetal bovine serum.
Downstream tips
Remaining cells were stained for FACS with anti-CD14, -16, -HLA-DR, and lineage exclusion markers as described for phenotyping above.
Downstream tips
Purity was assessed for all cells isolated by positive and negative selection.
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