Related Experiment Video
Updated: Feb 23, 2026
Isolation of Normal and Cancer-associated Fibroblasts from Fresh Tissues by Fluorescence Activated Cell Sorting FACS
Published on: January 14, 2013
Isolation of Fibroblast-Activation Protein-Specific Cancer-Associated Fibroblasts
Yingying Huang1, Sufang Zhou1, Yong Huang1
1National Center for International Research of Biological Targeting Diagnosis and Therapy, Guangxi Key Laboratory of Biological Targeting Diagnosis and Therapy Research, Collaborative Innovation Center for Targeting Tumor Diagnosis and Therapy, Guangxi Medical University, Nanning, Guangxi, China.
This study presents a new, gentle method to isolate cancer-associated fibroblasts (CAFs) using FAP magnetic beads. Twice collagenase IV digestion yielded more pure FAP+ CAFs than once, proving effective for cancer research.
Area of Science:
- Oncology
- Cell Biology
- Biotechnology
Background:
- Cancer-associated fibroblasts (CAFs) play a crucial role in tumor microenvironment.
- Efficient isolation of pure CAFs is essential for studying their function.
- Current methods may lack gentleness or efficiency.
Purpose of the Study:
- To develop a gentle and efficient method for purifying fibroblast-activation protein positive (FAP+) cancer-associated fibroblasts (CAFs) from tumor tissues.
- To optimize collagenase IV digestion for CAF isolation.
- To validate the purity and characteristics of isolated CAFs.
Main Methods:
- Tumor tissues from HepG2-bearing BALB/c-Nude mice were minced and digested.
- Cells were treated with trypsin and collagenase type IV (once, twice, or thrice).
- Purification was performed using FAP magnetic beads; FAP expression was analyzed.
Main Results:
- Twice collagenase IV digestion yielded significantly more cells (5.99 × 10^4) than once (2.58 × 10^4).
- The percentage of FAP+ CAFs was higher with twice digestion (38.5%) compared to once (20.0%).
- Isolated CAFs showed distinct FAP expression compared to normal fibroblasts and maintained CAF features.
Conclusions:
- A gentle and efficient method for FAP+ CAF purification was successfully developed.
- Optimized collagenase IV digestion (twice) enhances CAF yield and purity.
- The isolated CAFs are of high purity, in good condition, and exhibit characteristic CAF features.
