Source: Mac-Daniel, L., et al. Myeloid Cell Isolation from Mouse Skin and Draining Lymph Node Following Intradermal Immunization with Live Attenuated Plasmodium Sporozoites. J. Vis. Exp. (2016).
This video demonstrates an assay for immunizing a mouse model with live attenuated sporozoites. The radiation-attenuated Plasmodium sporozoites are injected into the dorsal side of the mouse's ear. After injections at multiple locations on both ears, the mouse is allowed to recover.
All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board.
1. Materials and Reagents
2. Radiation-attenuated Sporozoite Isolation from Mosquito Salivary Glands
3. Injection of Sporozoites into the Dermal Layer of the Ear
Figure 1: Intradermal Injection of Sporozoites into the Ear of Mouse. (A) Image showing intradermal injection of sporozoites in the ear pinna of an anesthetized mouse. The ventral side of the ear is stabilized with clear tape previously placed under a dissecting microscope. The needle is carefully inserted on the dorsal side of the ear, beneath the epidermis, with the bevel up. (B–C) Pictures showing the dorsal side of the ear pinna prior (B) and after (C) intradermal injection of 0.1-0.2 μl of parasite suspension. A characteristic papule (black arrow) is observable at the injection site at the end of the operation.
Figure 2: Imaging of Sporozoites Deposited in the Ear Dermis of Mouse. (A) Fluorescence microscopy showing RAS migrating from the injection site (indicated by the dashed lines) in the mouse skin 15 min post-injection (Scale bar = 60 μm; 10X magnification). The path of sporozoites is represented by the maximum intensity projection of the fluorescent signal. Green and red fluorescence respectively show parasite position in the skin at the beginning and after 10 sec of acquisition. (B) Higher magnification of RAS (green) injected in the skin (Scale bar = 20 μm; 25X magnification).
The authors have nothing to disclose.
Ketamine: Imalgene® 1000 | Merial | – | – |
Xylazine: Rompun® 2% | Bayer | – | – |
NanoFil syringe + 35 gauge needle | World Precision Instruments | – | – |
Omnican® 50 Insulin syringe 0,5 ml/50 I.U. | B. Braun Medical | 9151125 | – |
MultiwellTM 6 well tissue culture plate – Flat Bottom | BD Falcon | 353046 | – |
70 µm cell strainer | BD Falcon | 352350 | – |
2 ml syringe | Terumo | SS-02S | – |
BLUE MAXTM 15ml Polypropylene conical tube | BD Falcon | 352097 | – |
BLUE MAXTM 50ml Polypropylene conical tube | BD Falcon | 352098 | – |
5ml Polystyrene Round-Bottom Tube with 35µm Cell-Strainer Cap | BD Falcon | 352235 | – |
DPBS 1X Cacl2– and MgCl2-free | Life Technologies | 14190-094 | – |
DMEM 1X + GlutaMAXTM | Life Technologies | 31966-021 | – |
Deoxyribonuclease I from bovine pancreas, type IV | Sigma-Aldrich | D5025 | 50 µg/ml |
Female C57BL/6JRj mice (7-week-old) | Janvier Laboratories | – | – |
.