Mouse Lung Preparation for Flow Cytometry: Generating Cell Suspension from Lung Tissue for Flow Cytometric Analysis
Published: April 30, 2023
Abstract
Source: Moll, H. P., et al. Orthotopic Transplantation of Syngeneic Lung Adenocarcinoma Cells to Study PD-L1 Expression. J. Vis. Exp. (2019).
This video describes the preparation of mouse lung cell suspension for flow cytometric analysis to study PD-L1 expression of lung adenocarcinoma cells.
Protocol
All procedures involving animals have been reviewed by the local institutional animal care committee and the JoVE veterinary review board. 1. Lung Preparation for Flow Cytometry At the desired experimental endpoint, sedate the mouse by a subcutaneous injection of a mixture of ketamine (100 mg/kg of body weight) and xylazine (10 mg/kg of body weight) and euthanize it by cervical dislocation. Soak the carcass in 70% ethanol and secure the mouse on a dissect…
開示
The authors have nothing to disclose.
Materials
| Mouse lung adenocarcinoma cell line | isolated in-house | ||
| C57Bl/6 mice | F1 of the cross of the two backgrounds may be used (8-12 weeks) | ||
| 129S mice | |||
| RPMI 1640 Medium | Life Technologies | 11544446 | |
| Fetal Calf Serum | Life Technologies | 11573397 | |
| Penicillin/Streptomycin Solution | Life Technologies | 11548876 | |
| L-Glutamine | Life Technologies | 11539876 | |
| Ketasol (100 mg/mL Ketamine) | Ogris Pharma | 8-00173 | |
| Xylasol (20 mg/mL Xylazine) | Ogris Pharma | 8-00178 | |
| Blunt forceps | Roboz | RS8260 | |
| Student Iris Scissors | Fine Science Tools | 91460-11 | |
| DNase I (RNase-Free) | New England Biolabs | M0303S | |
| Collagenase Type I | Life Technologies | 17100017 | |
| ACK Lysing Buffer | Lonza | 10-548E |
