A Technique to Detect Extrapulmonary Tuberculosis via Superparamagnetic Nanoprobes

Abstract

Source: Lee, C., et al. Synthesis, Characterization, and Application of Superparamagnetic Iron Oxide Nanoprobes for Extrapulmonary Tuberculosis Detection. J. Vis. Exp. (2020).

This video demonstrates a technique for the detection of extrapulmonary tuberculosis. Superparamagnetic iron oxide (SPIO) nanoprobes, conjugated to Mycobacterium-specific antibodies, are injected into a mouse previously infected with Mycobacterium bovis BCG, labeling the granuloma and providing a better imaging contrast for its detection.

Protocol

All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board.

1. SPIO-MtbsAb synthesis

1. Synthesize SPIO-conjugated 2,2'-(ethylenedioxy)bisethylamine (EDBE) using previously reported methods.
2. Synthesize SPIO-EDBE-succinic anhydride (SA).
   1. Stir an alkaline solution (5 M NaOH; 10 mL)) of SPIO-EDBE and SA (1 g; 10 µmol) at room temperature for 24 h.
   2. Dialyze the solution with 20 changes of 2 L of distilled water using molecular porous membrane tubing (12,000-14,000 MW cutoff). 6 h for each change.

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
(benzotriazol-1-yloxy) tripyrrolidinophosphonium hexafluorophosphate	Sigma-Aldrich
1-hydroxybenzotriazole	Sigma-Aldrich
epichlorohydrin, 2,2'-(ethylenedioxy)bis(ethylamine)	Sigma-Aldrich
Human monocytic THP-1
M. bovis BCG	Pasteur Mérieux	Connaught strain; ImmuCyst Aventis
MRI	GE medical Systems	3.0-T, Signa
NMR relaxometer	Bruker	NMS-120 Minispec
SPECTRUM molecular porous membrane tubing, 12,000 -14,000 MW cut off	Spectrum Laboratories Inc
TB surface antibody- Polyclonal Antibody to Mtb	Acris Antibodies GmbH	BP2027
transmission electron microscope	JEOL	JEM-2000 EX II