Constructing a Humanized Mouse Model with Engineered Immunity against HIV
Abstract
Source: Zhen, A., et al. Stem-cell Based Engineered Immunity Against HIV Infection in the Humanized Mouse Model. J. Vis. Exp. (2016).
The video demonstrates a technique to generate a humanized mouse model with stem-cell-based engineered immunity against the human immunodeficiency virus (HIV). Human fetal thymus tissue and engineered hematopoietic stem cells (HSCs) are implanted into the kidney capsule of a severely immunocompromised mouse. The HSCs, which express chimeric antigen receptors (CARs) against HIV, are mixed in a gelatinous protein mixture that promotes their co-localization with thymus tissue. Post-procedure, the HSCs differentiate into various human immune cells, resulting in the development of a humanized mouse model with a functional human immune system against HIV.
Protocol
Offenlegungen
The authors have nothing to disclose.
Materials
| CD34 microbead kit | miltenyi | 130-046-702 | For sorting human CD34+ progenitor cells |
| Bambanker | Wako | 302-14681 | for freezing cells |
| QIAamp Viral RNA kit | Qiagen | 52904 | For measuring viral load in the serum |
| MACSQuant Flow Cytometer | Miltenyi | For flow analysis | |
| BD LSRFortessa™ | BD biosciences | For flow analysis | |
| Hyaluronidase | Sigma | H6254-500MG | For tissue digestion |
| Deoxyribonuclease I | Worthington | LS002006 | for tissue digestion |
| Collagenase | Life technology | 17104-019 | for tissue digestion |
| CFX Real time PCR detection system | Biorad | For measuring viral load and gene expression | |
| Mice, strain NOD.Cg-Prkdcscid Il2rgtm1Wjl/SzJ | The Jackson Laboratory | 5557 | For constructing the humanized mice |
| Penicillin Streptomycin (Pen Strep) | Thermo Fisher Scientific | 10378016 | For culturing cells |
| piperacillin/tazobactam | Pfizer | Zosyn | Anti-fungal |
| Amphotericin B (Fungizone antimycotic) | Thermo Fisher Scientific | 15290-018 | Anti-fungal |
| AUTOCLIP Wound Clips, 9 mm – 1000 units | Becton Dickinson | 427631 | For surgery |
| Sterile Poly-Reinforced Aurora Surgical Gowns, 30 per case | Medline | DYNJP2707 | For surgery |
| Sutures, 4-0, vicryl | Owens and Minor | 23000J304H | For surgery |
| Alcohol prep pads | Owens and Minor | 3583006818 | For surgery |
| Gloves, surgical, 6 1/2 | Owens and Minor | 4075711102 | For surgery |
| Yssel's Serum-Free T-Cell Medium | Gemini Bio-products | 400-102 | For CD34+ cell transduction |
| Human Serum Albumin | Sigma-Aldrich | A9511 | For CD34+ cell transduction |
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