二次谐波在生物医学成像中的应用

张子一; 王明雪; 刘志贺; 房晓峰; 吴长锋

doi:doi:10.3788/CJL202047.0207008

中国激光, 2020, 47 (2): 0207008, 网络出版: 2020-02-21

二次谐波在生物医学成像中的应用下载： 3370次特邀综述

Application of Second Harmonic Generation in Biomedical Imaging

张子一 ^1,2王明雪 ¹刘志贺 ¹房晓峰 ¹吴长锋 ^1,*

作者单位

¹ 南方科技大学生物医学工程系, 广东深圳 518000

² 哈尔滨工业大学生命科学与技术学院, 黑龙江哈尔滨 150000

图 & 表

图 1. 单光子荧光、双光子荧光和二次谐波的原理示意图。(a)单光子荧光和双光子荧光;(b)二次谐波产生^[4]

Fig. 1. Principles of single-photon excited fluorescence (SPEF), TPEF and SHG. (a) Single-photon excited fluorescence, two-photon excited fluorescence; (b) second harmonic generation^[4]

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图 2. SHG显微镜装置示意图及小鼠皮肤成像图^[13]

Fig. 2. SHG imaging schematic and example image of mouse skin^[13]

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图 3. 纤维状的胶原蛋白产生SHG信号的原理。(a)(b)胶原纤维作为偶极子;(c)不同偏振方向产生的SHG信号量^[13]

Fig. 3. SHG signaling generation principle for collagen fibers. (a)(b) Collagen fibers acting as dipoles; (c) the amount of SHG signal produced for different polarization orientations^[13]

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图 4. 将BaTiO₃纳米晶注射到单细胞阶段的斑马鱼胚胎中,持续观测发育过程中的SHG成像^[7]。(a)纵向检测(epi)斑马鱼躯干,SHG探针发出明亮、无背景噪音的信号;(b) SHG纳米探针靶向特异性

Fig. 4. BaTiO₃ nanocrystals were injected into zebrafish embryos at the single-cell stage for continuous SHG imaging during development^[7]. (a) SHG nanoprobes provide strong signal with superior signal-to-noise when zebrafish body is detected in epidirection; (b) SHG nanoprobe targeting specificity

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图 5. 在伤口愈合过程中,追踪内含有B-GQDs探针的小鼠间充质干细胞(mNSCs)^[36]

Fig. 5. Imaging and tracking mMSCs internalized with B-GQDs in wound healing in vivo over time^[36]

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图 6. SHG显微镜对微管网络的可视化结果。(a)微管SHG信号强度对激光偏振方向敏感;(b) SHG信号为窄峰;(c)(d) SHG信号来源于微管极化结构^[59]

Fig. 6. Visualization results of microtubule network by SHG microscope. (a) SHG signal intensity of microtubule is sensitive to laser polarization; (b) SHG signal is narrow peak; (c)(d) SHG signal comes from microtubule polarization structure^[59]

下载图片查看原文

图 7. 动作电位的SHG记录。(a)用SHG测体外培养神经元膜电位的线性扫描记录;(b)沿神经元轴突动作电位的SHG记录^[60]

Fig. 7. SHG recording of action potentials. (a) Line-scan recording of membrane potential with SHG in cultured neurons; (b) SHG recording of action potentials along neuron neurite^[60]

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图 8. 使用二次谐波特异性染料的多模态双光子成像。(a)染料Ap3和FM4-64的化学结构;(b)神经元动作电位的SHG信号;(c) FM4-64和Ap3标记CHO细胞的SHG和TPF信号;(d)(e)细胞结构的多模态双光子成像;(d)使用FM4-64前后TPF信号的强度分布;(e)使用FM4-64的合并图像^[35]

Fig. 8. Multimodal two-photon imaging using a second harmonic generation-specific dye. (a) Chemical structures of Ap3 and FM4-64; (b) SHG signal changes on action potential in neurons; (c) SHG and TPF signals obtained from cultured CHO cells loaded with FM4-64 and Ap3; (d)(e) multimodal two-photon imaging of cellular structures; (d) intensity profiles of TPF signals before and after the application of FM4-64; (e) merged images after the application of FM4-64^{[35
下载图片查看原文}

表 1二次谐波生物医学成像探针

Table1. SHG probes for biomedical imaging

SHG probe	Type of materials	λ_max/nm (solvent)	β /(10^-28 esu)
KNb $\begin{matrix} {O_{3}}^{[15]} \end{matrix}$	Inorganic nanocrystal	502	7.2×10⁴
BaTi $\begin{matrix} {O_{3}}^{[16 - 18]} \end{matrix}$	Inorganic nanocrystal	425	11.0×10⁴
LiNb $\begin{matrix} {O_{3}}^{[25 - 26]} \end{matrix}$	Inorganic nanocrystal	—	4.8 ×10⁴
ZnO^[5,15,27]	Inorganic nanocrystal	502	9.6×10⁴
Fe(IO₃ $\begin{matrix} {)_{3}}^{[28 - 29]} \end{matrix}$	Inorganic nanocrystal	—	—
BiFe $\begin{matrix} {O_{3}}^{[19]} \end{matrix}$	Inorganic nanocrystal	—	1.2×10⁶
DANS^[30]	Organic dye	430 (CHCl₃)	7×10^-1
DIA^[30]	Organic dye	590 (CHCl₃)	1.9
RH237^[31]	Organic dye	500 (MeCN)	9.0
FM4-64^[32-33]	Organic dye	500 (MeCN)	1.1×10
di-4-ANEPPS^[34]	Organic dye	542 (CHCl₃)	—
JPW1259^[34]	Organic dye	542 (CHCl₃)	—
Ap3^[35]	Organic dye	—	—
B-GQD^[36]	Graphene quantum dots	—	—

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表 2生物医学成像探针的特点

Table2. Features of biomedical imaging probes

Biomedical imaging probe	No blinking	No bleaching	Non-absorptive	Deepspectrum	Narrow spectrum(<10 nm)	Geneticallyencoded
SHG nanoprobes^[4]	√	√	√	√	√
Fluorescent dyes^[37]
Fluorescent proteins^[46]						√
UC nanoparticles^[38-40,45]	√	√
SERS nanoparticles^[41-44]		√		√	√

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张子一, 王明雪, 刘志贺, 房晓峰, 吴长锋. 二次谐波在生物医学成像中的应用[J]. 中国激光, 2020, 47(2): 0207008. Zhang Ziyi, Wang Mingxue, Liu Zhihe, Fang Xiaofeng, Wu Changfeng. Application of Second Harmonic Generation in Biomedical Imaging[J]. Chinese Journal of Lasers, 2020, 47(2): 0207008.

二次谐波在生物医学成像中的应用下载： 3370次特邀综述

图 1. 单光子荧光、双光子荧光和二次谐波的原理示意图。(a)单光子荧光和双光子荧光;(b)二次谐波产生^[4]

Fig. 1. Principles of single-photon excited fluorescence (SPEF), TPEF and SHG. (a) Single-photon excited fluorescence, two-photon excited fluorescence; (b) second harmonic generation^[4]

图 2. SHG显微镜装置示意图及小鼠皮肤成像图^[13]

Fig. 2. SHG imaging schematic and example image of mouse skin^[13]

图 3. 纤维状的胶原蛋白产生SHG信号的原理。(a)(b)胶原纤维作为偶极子;(c)不同偏振方向产生的SHG信号量^[13]

Fig. 3. SHG signaling generation principle for collagen fibers. (a)(b) Collagen fibers acting as dipoles; (c) the amount of SHG signal produced for different polarization orientations^[13]

图 4. 将BaTiO₃纳米晶注射到单细胞阶段的斑马鱼胚胎中,持续观测发育过程中的SHG成像^[7]。(a)纵向检测(epi)斑马鱼躯干,SHG探针发出明亮、无背景噪音的信号;(b) SHG纳米探针靶向特异性

图 5. 在伤口愈合过程中,追踪内含有B-GQDs探针的小鼠间充质干细胞(mNSCs)^[36]

Fig. 5. Imaging and tracking mMSCs internalized with B-GQDs in wound healing in vivo over time^[36]

图 6. SHG显微镜对微管网络的可视化结果。(a)微管SHG信号强度对激光偏振方向敏感;(b) SHG信号为窄峰;(c)(d) SHG信号来源于微管极化结构^[59]

Fig. 6. Visualization results of microtubule network by SHG microscope. (a) SHG signal intensity of microtubule is sensitive to laser polarization; (b) SHG signal is narrow peak; (c)(d) SHG signal comes from microtubule polarization structure^[59]

图 7. 动作电位的SHG记录。(a)用SHG测体外培养神经元膜电位的线性扫描记录;(b)沿神经元轴突动作电位的SHG记录^[60]

Fig. 7. SHG recording of action potentials. (a) Line-scan recording of membrane potential with SHG in cultured neurons; (b) SHG recording of action potentials along neuron neurite^[60]

表 1二次谐波生物医学成像探针

Table1. SHG probes for biomedical imaging

表 2生物医学成像探针的特点

Table2. Features of biomedical imaging probes

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二次谐波在生物医学成像中的应用 下载： 3370次特邀综述

图 1. 单光子荧光、双光子荧光和二次谐波的原理示意图。(a)单光子荧光和双光子荧光;(b)二次谐波产生[4]

Fig. 1. Principles of single-photon excited fluorescence (SPEF), TPEF and SHG. (a) Single-photon excited fluorescence, two-photon excited fluorescence; (b) second harmonic generation[4]

图 2. SHG显微镜装置示意图及小鼠皮肤成像图[13]

Fig. 2. SHG imaging schematic and example image of mouse skin[13]

图 3. 纤维状的胶原蛋白产生SHG信号的原理。(a)(b)胶原纤维作为偶极子;(c)不同偏振方向产生的SHG信号量[13]

Fig. 3. SHG signaling generation principle for collagen fibers. (a)(b) Collagen fibers acting as dipoles; (c) the amount of SHG signal produced for different polarization orientations[13]

图 4. 将BaTiO3纳米晶注射到单细胞阶段的斑马鱼胚胎中,持续观测发育过程中的SHG成像[7]。(a)纵向检测(epi)斑马鱼躯干,SHG探针发出明亮、无背景噪音的信号;(b) SHG纳米探针靶向特异性

Fig. 4. BaTiO3 nanocrystals were injected into zebrafish embryos at the single-cell stage for continuous SHG imaging during development[7]. (a) SHG nanoprobes provide strong signal with superior signal-to-noise when zebrafish body is detected in epidirection; (b) SHG nanoprobe targeting specificity

图 5. 在伤口愈合过程中,追踪内含有B-GQDs探针的小鼠间充质干细胞(mNSCs)[36]

Fig. 5. Imaging and tracking mMSCs internalized with B-GQDs in wound healing in vivo over time[36]

图 6. SHG显微镜对微管网络的可视化结果。(a)微管SHG信号强度对激光偏振方向敏感;(b) SHG信号为窄峰;(c)(d) SHG信号来源于微管极化结构[59]

Fig. 6. Visualization results of microtubule network by SHG microscope. (a) SHG signal intensity of microtubule is sensitive to laser polarization; (b) SHG signal is narrow peak; (c)(d) SHG signal comes from microtubule polarization structure[59]

图 7. 动作电位的SHG记录。(a)用SHG测体外培养神经元膜电位的线性扫描记录;(b)沿神经元轴突动作电位的SHG记录[60]

Fig. 7. SHG recording of action potentials. (a) Line-scan recording of membrane potential with SHG in cultured neurons; (b) SHG recording of action potentials along neuron neurite[60]

表 1二次谐波生物医学成像探针

Table1. SHG probes for biomedical imaging

表 2生物医学成像探针的特点

Table2. Features of biomedical imaging probes

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二次谐波在生物医学成像中的应用下载： 3370次特邀综述

图 1. 单光子荧光、双光子荧光和二次谐波的原理示意图。(a)单光子荧光和双光子荧光;(b)二次谐波产生^[4]

Fig. 1. Principles of single-photon excited fluorescence (SPEF), TPEF and SHG. (a) Single-photon excited fluorescence, two-photon excited fluorescence; (b) second harmonic generation^[4]

图 2. SHG显微镜装置示意图及小鼠皮肤成像图^[13]

Fig. 2. SHG imaging schematic and example image of mouse skin^[13]

图 3. 纤维状的胶原蛋白产生SHG信号的原理。(a)(b)胶原纤维作为偶极子;(c)不同偏振方向产生的SHG信号量^[13]

Fig. 3. SHG signaling generation principle for collagen fibers. (a)(b) Collagen fibers acting as dipoles; (c) the amount of SHG signal produced for different polarization orientations^[13]

图 4. 将BaTiO₃纳米晶注射到单细胞阶段的斑马鱼胚胎中,持续观测发育过程中的SHG成像^[7]。(a)纵向检测(epi)斑马鱼躯干,SHG探针发出明亮、无背景噪音的信号;(b) SHG纳米探针靶向特异性

图 5. 在伤口愈合过程中,追踪内含有B-GQDs探针的小鼠间充质干细胞(mNSCs)^[36]

Fig. 5. Imaging and tracking mMSCs internalized with B-GQDs in wound healing in vivo over time^[36]

图 6. SHG显微镜对微管网络的可视化结果。(a)微管SHG信号强度对激光偏振方向敏感;(b) SHG信号为窄峰;(c)(d) SHG信号来源于微管极化结构^[59]

Fig. 6. Visualization results of microtubule network by SHG microscope. (a) SHG signal intensity of microtubule is sensitive to laser polarization; (b) SHG signal is narrow peak; (c)(d) SHG signal comes from microtubule polarization structure^[59]

图 7. 动作电位的SHG记录。(a)用SHG测体外培养神经元膜电位的线性扫描记录;(b)沿神经元轴突动作电位的SHG记录^[60]

Fig. 7. SHG recording of action potentials. (a) Line-scan recording of membrane potential with SHG in cultured neurons; (b) SHG recording of action potentials along neuron neurite^[60]