Isolating RPE and IPE Cells from Bovine Eye: A Procedure to Harvest and Culture Bovine Primary Pigment Epithelial Cells
Published: April 30, 2023
Abstract
Source: Bascuas, T. et al., Isolation, Culture, and Genetic Engineering of Mammalian Primary Pigment Epithelial Cells for Non-Viral Gene Therapy. J. Vis. Exp. (2021).
This video demonstrates the procedure to isolate the retinal pigment epithelial, or RPE, cells and iris pigment epithelial, or IPE, cells from harvested bovine eyes. The isolated cells can be cultured to study their role in the treatment of ocular diseases.
Protocol
All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board. 1. Isolation of bovine PE cells Use curved scissors and Colibri forceps to enucleate the eyes after euthanizing the animal. Clean the remaining muscle tissue and skin from the eyes using scissors and forceps (non-sterile). NOTE: The size of the scissors and forceps used for the enucleation and cleaning of the eyes d…
Representative Results
Figure 1: Instruments used for PE isolation depending on the species. (A) Non-sterile instruments used for the enucleation of the eyes and cleaning of remaining muscle tissue and skin. Set 1 is used for rat, mouse, and rabbit, and set 2 is used for pig and cattle eyes. (B) Sterile instruments used for PE isolation. Note the different size of scissors and forceps …
Divulgations
The authors have nothing to disclose.
Materials
| 6-well plate | Greiner | 7657160 | |
| Betadine | Mundipharma | ||
| Colibri forceps (sterile) | |||
| Forceps (different size) (sterile) | |||
| Gauze compress | PROMEDICAL AG | 25403 | |
| Neubauer chamber | Marienfeld-superior | 640010 | |
| Pasteur pipette (fire-polish) | Witeg | 4100150 | |
| PBS 1X | Sigma-Aldrich | D8537 | |
| Penicillin/Streptomycin | Sigma-Aldrich | P0781-100 | |
| Petri dish | ThermoFisher Scientific | 150288 | |
| DMEM/Ham`s F12 | Sigma-Aldrich | D8062 | |
| FBS | Brunschwig | P40-37500 | |
| Scalpel no. 10 | Swann-Morton | 501 | |
| Scalpel no. 11 | Swann-Morton | 503 | |
| Sharp-sharp tip curved Extra Fine Bonn Scissors (sterile) | |||
| Sharp-sharp tip straight Extra Fine Bonn Scissors (sterile) | |||
| Trypsin 0.25% | ThermoFisher Scientific | 25050014 | |
| Vannas spring scissors curved (sterile) |
Tags
Citer Cet Article
Isolating RPE and IPE Cells from Bovine Eye: A Procedure to Harvest and Culture Bovine Primary Pigment Epithelial Cells. J. Vis. Exp. (Pending Publication), e20829, doi: (2023).