The adipose tissue stromal-vascular fraction was obtained as described previously (3 (link)) and resuspended in PBS supplemented with 2% FCS. Surface markers were stained with saturating amounts of the following fluorescent-labeled monoclonal antibodies (BioLegend) for flow cytometry: CD11c (clone N418), CD11b (clone M1/70), F4/80 (clone CI:A3–1), CD206 (clone C068C2), and CD45 (BD Horizon clone 30-F11). For intracellular cytokine staining, cells were stimulated in vitro for 5 h at 37°C in 5% CO2 with a leukocyte activation cocktail (BD Biosciences) and stained with anti–tumor necrosis factor (TNF) antibody (clone MP6-XT22; BioLegend) as described previously (3 (link)). Cells were acquired on a specially ordered five-laser LSR II Flow Cytometer (BD Biosciences) at the Beth Israel Deaconess Medical Center Flow Cytometry Core and analyzed with FlowJo software (Tree Star).
Protocol detail
Isolation and Analysis of Adipose-Derived Immune Cells
Partial Protocol Preview
This section provides a glimpse into the protocol.
The remaining content is hidden due to licensing restrictions, but the full text is available at the following link:
Access Free Full Text.
Adipose stromal vascular fraction
Anti antibody
Cd11b
Cells
Clone
Cytokine
Flow cytometry
Intracellular
Leukocyte
Monoclonal antibodies
Tissue
Tree
Tumor necrosis factor
Corresponding Organization : Harvard University
Other organizations : Ionis Pharmaceuticals (United States)
Top 5 similar protocols
Protocol cited in 1 other protocol
Variable analysis
independent variables
- Stimulation with a leukocyte activation cocktail (BD Biosciences)
- Expression of tumor necrosis factor (TNF)
- Expression of surface markers: CD11c, CD11b, F4/80, CD206, CD45
- Adipose tissue stromal-vascular fraction obtained as described previously (3 (link))
- Resuspension of cells in PBS supplemented with 2% FCS
- Staining of surface markers with saturating amounts of fluorescent-labeled monoclonal antibodies (BioLegend)
- None specified
- None specified
Annotations
Based on most similar protocols
Etiam vel ipsum. Morbi facilisis vestibulum nisl. Praesent cursus laoreet felis. Integer adipiscing pretium orci. Nulla facilisi. Quisque posuere bibendum purus. Nulla quam mauris, cursus eget, convallis ac, molestie non, enim. Aliquam congue. Quisque sagittis nonummy sapien. Proin molestie sem vitae urna. Maecenas lorem.
Sign up for free or login to display all annotations
As authors may omit details in methods from publication, our AI will look for missing critical information across the 5 most similar protocols.
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!