A Bright NIR-II Fluorescence Probe for Vascular and Tumor Imaging

Summary

The present protocol describes a detailed, real-time NIR-II fluorescence imaging operation of a mouse using a NIR-II optics imaging device.

Abstract

As an emerging imaging technology, near-infrared II (NIR-II, 1000-1700 nm) fluorescence imaging has significant potential in the biomedical field, owing to its high sensitivity, deep tissue penetration, and superior imaging with spatial and temporal resolution. However, the method to facilitate the implementation of NIR-II fluorescence imaging for some urgently needed fields, such as medical science and pharmacy, has puzzled relevant researchers. This protocol describes in detail the construction and bioimaging applications of a NIR-II fluorescence molecular probe, HLY1, with a D-A-D (donor-acceptor-donor) skeleton. HLY1 showed good optical properties and biocompatibility. Furthermore, NIR-II vascular and tumor imaging in mice was performed using a NIR-II optics imaging device. Real-time high-resolution NIR-II fluorescence images were acquired to guide the detection of tumors and vascular diseases. From probe preparation to data acquisition, the imaging quality is greatly improved, and the authenticity of the NIR-II molecular probes for data recording in intravital imaging is ensured.

Introduction

Fluorescence imaging is the commonly used molecular imaging tool in basic research, and is also often used to guide surgical tumor resection in clinics¹. The essential principle of fluorescence imaging is to employ a camera to receive fluorescence emitted by a laser after the irradiation of samples (tissues, organs, etc.)². The process is completed within a few milliseconds³. The fluorescence imaging wavelengths can be divided into ultraviolet (200-400 nm), visible region (400-700 nm), near-infrared I (NIR-I, 700-900 nm), and near-infrared II (NIR-II, 1000-1700 nm)^4,

Protocol

Animal experiments for NIR-II imaging studies were conducted at the Animal Experiment Center of Wuhan University, which has been awarded the International Association for Experimental Animal Care (AALAC). All animal studies were conducted following the China Animal Welfare Commission Guidelines for the Care and Use of Experimental Animals and approved by the Animal Care and Use Committee (IACUC) of the Animal Experimental Center of Wuhan University.

Female BALB/c nude mice (~20 g) at 6 weeks of age were used for the present study.

1. NIR-II imaging preparation

1. Place commercially avail....

Results

The fluorescent intensity and brightness of water-suspensible HLY1 dots were determined by an NIR-II imaging instrument. The fluorescent intensity of HLY1 in the 90% f_wTHF/H₂O mixture was five times that in the THF solution, which indicated a prominent AIE feature of HLY1 (Figure 1B). Moreover, HLY1 dots emitted strong fluorescent signals under a 1,500 nm LP filter, showing that HLY1 dots can be used for NIR-IIb imaging (Figure 1D).......

Discussion

NIR-I fluorescent imaging can be used to some extent for tumor and vascular imaging, but due to the limited maximum emission wavelength of NIR-I fluorophores (<900 nm), it results in poor tissue penetration and tumor signal background ratio^33,34. Poor and low imaging resolution may cause a deviation between the outcome of the imaging feedback treatment and the actual therapeutic effect. In addition, most NIR-I fluorophores have poor optical stability and extr.......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
Anhydrous pyridine	Perimed	110-86-1
Anhydrous sodium sulfate	China national medicines Co.,Ltd	SY006376
Black cardboard	Suzhou Yingrui Optical Technology Co., Ltd	AO00158
Column chromatography	Energy Chemical	E080498
Diphenylphosphine palladium dichloride	Sigma-Aldrich	B2161-1g
DSPE-PEG2000	Ponsure	PS-E1
Dulbecco's modified eagle medium	Gibco	8121587
EGTA	Biofroxx	EZ6789D115
Fetal bovine serum	Gibco	2166090RP
Isoflurane	GLPBIO	GC45487-1
K2CO3	Macklin	P816305-5g
N. N '- dimethylformamide	China national medicines Co.,Ltd	02-12-1968
NIR-II imaging instrument	Suzhou Yingrui Optical Technology Co., Ltd	16011109
N-sulfenanilide	Enerry chemical	1250030-5g
PdCl2(dppf)2CH2Cl2	TCI	B2064-1g
penicillin-streptomycin	Gibco	15140-122
Tetrahydrofuran	China national medicines Co.,Ltd	M005197
Tetratriphenylphosphine palladium	Immochem	1021232-5g
Tetratriphenylphosphine palladium	Sigma-Aldrich	1021232-5g
Tributyltin chloride	Immochem	QH004335
Trimethylchlorosilane	China national medicines Co.,Ltd	40060560