For the analysis of costimulatory molecule expression, cells were incubated with AQUA LIVE/DEAD fluorescent dye (0.5 µl) (Invitrogen) for 30 min. Cells were subsequently stained with 0.3 µg/ml of anti-CD11c (HL3), 1.25 µg/ml of anti-CD86 (24F) both obtained from BD Biosciences and 2.5 µg/ml of anti-CD40 (3.23) from eBiosciences. Mitochondrial ROS were measured in cells by MitoSOX (Invitrogen) staining (1 µM for 15 min at 37°C) (Nakahira et al., 2011 (link); Tal et al., 2009 (link)). Samples were acquired on BD FACSCanto II or Cyan using FACSDiva (BD) or Summit (Dako) software and the data analyzed using FlowJo software (Treestar).
Anti cd86 24f
Manufactured by BD
Anti-CD86 (24F) is a monoclonal antibody that binds to the CD86 antigen. CD86 is a costimulatory molecule expressed on antigen-presenting cells that plays a role in T cell activation. The antibody can be used in flow cytometry applications to identify and quantify CD86-expressing cells.
Automatically generated - may contain errors
2 protocols using anti cd86 24f
1
Costimulatory Molecule Expression and Mitochondrial ROS Analysis
2
Multicolor Flow Cytometry of Immune Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Cells were pre-incubated with rat serum (diluted 1:100 in PBS) for 10 min at 4°C to minimize nonspecific antibody binding. Cells were then suspended in FACS buffer (PBS containing 2% FCS and 0.05% NaN 2 ) and incubated with primary antibodies against the cell-surface markers: anti-CD45 (OX-1; BD Pharmingen TM ), anti-CD86 (24F, BD Pharmingen TM ), anti-CD163 (ED2, Bio-Rad), or anti-CCR2 (890231, R&D SYSTEMS) for 30 min at 4°C. 7-AAD was used to differentiate between live and dead cells. For intracellular staining to determine CD68 levels, cells were fixed after surface staining and then permeabilized using 0.3% Triton X-100 for 30 min, before anti-CD68 staining (ED1, Bio-Rad) for 30 min. Cell counts were determined using a BD FACS Canto II flow cytometer (BD Biosciences) and analyzed with FlowJo software (FlowJo, LLC, Ashland, OR). The gating strategy and control experiments are shown in Supplementary Fig. 3. The antibodies are shown in Supplementary table 1.
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!