A Murine Model of Primed Mycobacterial Uveitis to Study Post-Infectious Uveitis

Abstract

Source: John, S., et al. Primed Mycobacterial Uveitis (PMU) as a Model for Post-Infectious Uveitis. J. Vis. Exp. (2021).

This video demonstrates a method to generate a mouse model of primed mycobacterial uveitis (PMU) for studying post-infectious uveitis. Upon induction of PMU by intravitreal injection of heat-killed Mycobacterium tuberculosis in a mouse previously immunized with the same immunogen, the progression of chronic inflammation in the eye is analyzed using imaging and histopathological examination

Protocol

All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board. 1. Antigen preparation for subcutaneous injection Perform all procedures in this section inside a chemical fume hood to prevent inhalation or skin contact with the Mycobacterium tuberculosis (Mtb) H37Ra powder. Handle according to your institutional policies for Complete Freund's Adjuvant (typically BSL-1). Th…

Representative Results

Figure 1: Mouse intravitreal injection setup. The intravitreal injection is performed on the mouse eye using (A) a micro syringe pump controller connected to the (B) Micropump and (C) an injection syringe. The syringe is loaded and mounted on the (D) Micropump. The mouse head is positioned using (E) ear bars to e…

Materials

AK-FLUOR	Akorn Pharmaceuticals, IL, USA		10% Fluorescein sodium 100 mg/mL in 5 mL vial
AnaSed	Akorn Animal Health, IL, USA	NDC 59399-110-20	Xylazine 20 mg/mL
Betadine 5% Sterile Ophthalmic Prep Solution	Alcon, TX, USA	8007-1
B-D Precision Glide Needles -25 G	Becton, Dickinson and Company, NJ, USA	305122
B-D Precision Glide needle -30-G	Becton, Dickinson and Company, NJ, USA	305106
Bond MAX, Bond Rx	Leica Biosystems, IL,USA		Automated IHC staining system
Chloramphenicol ointment	Martindale Pharma, Romford, UK		1% w/w Chloramphenicol
EG1150H	Leica Biosystems, IL,USA		Tissue Embedding
Envisu R2300	Bioptigen/Leica		OCT Machine
Freund's Incomplete Adjuvant	BD Difco, NJ, USA	263910
GenTeal lubricant eye ointment	Alcon, TX, USA	—
GenTeal lubricant eye gel	Alcon, TX, USA	—
H37Ra lyophilized Mycobacteria extract	BD Difco, NJ, USA	231141
Hamilton RN Needle (33/12/2)S	Hamilton, Reno, NV	7803-05(33/12/2)	33 G
Hamilton syringe	Hamilton, Reno, NV	CAL7633-01	5 µL
Insulin needle	Exel International, USA	26029	1 mL
Isoflurane
Ketaset	Zoetis, USA	377341	Ketamine HCL 100 mg/mL
Microinjection Syringe Pump and Micro4Controller	World Precision Instruments, FL, USA	UMP3
Micron IV	Phoenix Research Laboratories, Pleasanton, CA		Alternative Imaging/OCT Machine
Nanofil 10 µL syringe	World Precision Instruments, FL, USA	NANOFIL
Nanofil Intraocular Injection Kit	World Precision Instruments, FL, USA	IO-KIT
Olympus SZX10	Olympus		Dissection scope
PBS	Gibco	14190
Phenylephrine Hydrochloride Ophthalmic Solution USP 2.5% Sterile 15 mL	Akorn Pharmaceuticals, IL, USA	17478020115
RM2255	Leica Biosystems, IL,USA		Tissue Sectioning
TB Syringe	Becton, Dickinson and Company, NJ, USA	309602	1 mL
Tetracaine 0.5%	Alcon, TX, USA	1041544
Tissue Tek VIP series	Sakura Finetek USA, Inc.,CA.		Histology Tissue Processing
Tropicamide 1%	Chauvin Pharmaceuticals, Romford, UK		Minims
Tylenol	Johnson & Johnson Consumer Inc, PA, USA	NDC 50580-614-01	Acetaminophen
Viscotears	Novartis Pharmaceuticals, Camberley, UK		Carbomer eye gel 0.2% w/w