Intracellular Analysis of NK Cell Activation
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.
Corresponding Organization : The Ohio State University Comprehensive Cancer Center – Arthur G. James Cancer Hospital and Richard J. Solove Research Institute
Other organizations : The Barbara Ann Karmanos Cancer Institute, OncoMed (United States), Wright State University, Grady Memorial Hospital
Variable analysis
- C-IgG– or F-IgG–FITC treatment of KB tumor cells
- Intracellular analysis of NK cell ERK phosphorylation following coculture with KB tumor cells
- NK cell activation assessed 48 hours post coculture via FACS analysis with various markers (CD56, CD158, CD57, NKG2A, CD56, CD3, CD16, NKG2D, CD69, KIR/NKAT2, CD94, NKp46/CD335)
- FR⁺/FITC⁺ populations in dissociated tumor cells from murine studies with C‐IgG– and F‐IgG–FITC
- Incubation time (30 min) for C-IgG– or F-IgG–FITC treatment of KB tumor cells
- Temperature (37°C) for C-IgG– or F-IgG–FITC treatment of KB tumor cells
- Washing with flow buffer (5% FBS in PBS) prior to fixation
- Fixation in 1% formalin in PBS
- Mounting on 25 × 75 × 1 mm slides (Erie Scientific, Portsmouth, NH)
- Staining with DAPI for nuclear identification (Vector Labs, Burlingame, CA)
- Murine tumor sections analyzed using Olympus Fluoview 1000 laser scanning confocal microscope
- Tumor cells dissociated and stained with FR‐α–APC (R&D Systems, Minneapolis, MN)
- FR⁺/FITC⁺ populations assessed using Becton Dickinson FACS Calibur flow cytometer (Becton‐Dickinson, San Jose, CA)
Annotations
Based on most similar protocols
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!