Immunoisolation of pancreatic epithelial cells from endoscopic ultrasound-guided fine needle aspirates with magnetic beads for downstream molecular application.

BACKGROUND

Evaluation of pancreatic mass is routinely performed by endoscopic ultrasound-guided fine needle aspiration (EUS-FNA). However, molecular analyses of the tumor cells on FNA samples are limited by the significant cellular heterogeneity. The goal of the current study is to evaluate a magnetic immunoconcentration technique in isolating pancreatic epithelial cells from needle aspirates, and to demonstrate that the isolated cells could be utilized for molecular analysis.

METHODS

Pancreatic EUS-FNA specimens were processed and stored at -80°C. Based on the cytopathological diagnosis, 17 adenocarcinoma, 3 lymphoproliferative, and 3 benign cases were retrieved from the collection for further analyses. Epithelial cells were isolated using antihuman epithelial cell specific antibody-bound magnetic beads, and the isolated cellular component was confirmed cytologically. Genomic DNA was extracted, quantitated, and evaluated with methylation-specific PCR for cyclin D2 in 8 adenocarcinoma cases.

RESULTS

After optimization, malignant epithelial cells were successfully isolated from all adenocarcinoma cases. Normal pancreatic ductal cells were isolated from three benign cases. No cells were retrieved after immunomagnetic isolation in all three lymphoproliferative cases.  DNA yields were 5-2646 ng, with a mean of 357 ng. Methylation-specific PCR for cyclin D2 on the 8 carcinoma cases showed methylated state at the promoter region, demonstrating feasible evaluation of the methylation status.

CONCLUSION

The magnetic immunoconcentration of pancreatic EUS-FNA specimen described here is a practical method of isolating pancreatic epithelial cells from needle aspirates. Isolated cells were sufficient for performing subsequent molecular analysis.