Induction of Graft Versus Host Disease in Mouse Models via Allogeneic Bone Marrow and Splenic Cell Transplantation
Published: October 31, 2023
Abstract
Source: Buchele, V., et al. Induction of Intestinal Graft-versus-host Disease and Its Mini-endoscopic Assessment in Live Mice. J. Vis. Exp. (2019).
This video demonstrates the induction of intestinal graft-versus-host disease, GvHD, in a lethally-irradiated recipient mouse using allogeneic hematopoietic stem cells and GvHD-inducing splenic alloreactive T lymphocytes. This protocol helps establish a severe acute intestinal GvHD model to understand the mechanism of the disease and develop innovative therapeutic options.
Protocol
All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board. 1. GvHD induction Day 0: Total body irradiation of the recipient mice Use female CD45.2+ H2kd+ BALB/c mice that are at least 10 weeks old as recipients. Weigh and record the weight of the recipient mice prior to GvHD induction. NOTE: Ensure t…
Divulgations
The authors have nothing to disclose.
Materials
| BIOBEAM 2000 Gamma Irradiator | Gamma-Service Meical GmbH | ||
| Phosphate-buffered saline (PBS ) | Sigma-Aldrich Co. LLC. | D8662-6x500ML | |
| Semken Forceps (lenght: 13 cm; serrated, curved tips) | Fine Science Tools | 11009-13 | |
| Hardened Fine Scissors (lenght: 8,5 cm; straight tips; cutting edge: 24 mm) | Fine Science Tools | 14090-09 | |
| RPMI-1640 Medium | Sigma-Aldrich Co. LLC. | R8758-500ML | |
| Hypodermic needle (26G) | B. Braun Melsungen AG | 4657683 | |
| 1 mL syringe | B. Braun Melsungen AG | 9166017V | |
| 50 mL tube | Sarstedt Ag & Co.KG | 6,25,47,254 | |
| Cell strainer with a 40 µm mesh screen | BD Falcon | 352340 | |
| Insulin syringe with sterile interior (30G) | BD | 324826 | |
| Pan T Cell Isolation Kit II, mouse | Miltenyi Biotec GmbH | 130-095-130 | magnet cell separation to isolate splenic T cells |
| CD90.2 MicroBeads, mouse | Miltenyi Biotec GmbH | 130-049-101 | magnet cell separation to isolate T cell-depleted bone marrow cells |
| Biopsy Forceps | KARL STORZ SE &Co KG | 61071 ZJ | part of the experimental setup for colonoscopy |
| Ethylenediaminetetraacetic acid disodium salt dihydrate (Na2EDTA) | Carl Roth GmbH & Co.KG | 8043.2 | ingredient of ACK lysing buffer |
| Alexa Fluor 700 anti-mouse CD45.2 Antibody (clone: 104; lot: B252126; RRID: AB_493731) | Biolegend | 109822 | |
| PE/Cy7 anti-mouse CD45.1 Antibody (clone: A20; lot: B217246; RRID: AB_1134168) | Biolegend | 110730 | |
| APC/Cy7 anti-mouse CD8a Antibody (clone: 53-6.7, lot: B247008; RRID: AB_312753) | Biolegend | 100714 | |
| Forane / Isofluran | AbbVie Inc. | B506 | |
| Filtrated bovine serum | Pan Biotec | P40-47500 | ingredient of FACS buffer |
Tags
Citer Cet Article
Induction of Graft Versus Host Disease in Mouse Models via Allogeneic Bone Marrow and Splenic Cell Transplantation. J. Vis. Exp. (Pending Publication), e21718, doi: (2023).