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Bioengineering

基于脉冲激光二极管的桌面光声层析成像在大鼠皮质血管内监测染料的洗入和洗脱

Published: May 30, 2019
doi:
10.3791/59764
, , , ,
1School of Chemical and Biomedical Engineering,Nanyang Technological University

Summary

基于平板脉冲激光二极管的桌面光声层析成像 (LDD-PAT) 系统被证明用于小动物皮质血管的高速动态体内成像。

Abstract

光声 (PA) 断层扫描 (pat) 成像是一种新兴的生物医学成像模式, 适用于各种临床前和临床应用。定制的基于圆形环阵列的传感器和传统的笨重 Nd: YAG/OPO 激光器抑制了 PAT 系统的转换到诊所。超紧凑型脉冲激光二极管 (Pld) 目前正被用作 PA 成像的近红外激发的替代来源。高速动态体内成像已被证明使用了紧凑型基于 pld 的桌面 PAT 系统 (PLD-PAT)。本文提出了一种利用桌面 PLD-PAT 系统进行动态脑成像的可视化实验方案。该协议描述了桌面 PLD-PAT 系统配置、脑血管成像动物的制备以及大鼠皮质血管中肌内腺绿色 (ICG) 染料吸收和清除过程的动态成像过程。

Introduction

光声计算机断层扫描 (pactse-pat) 是一种很有前途的非侵入性生物医学成像方式, 它结合了丰富的光学对比度和高超声分辨率1,2,3, 4,5. 当纳秒脉冲激光将能量沉积到任何生物组织内的吸光色相中时, 局部温度升高, 导致组织的热弹性膨胀和收缩, 从而产生压力波。这些压力波被称为超声波或光声 (PA) 波, 可以通过样品周围的超声波传感器检测到。利用各种重建算法6、7、8、9对检测到的 pa 信号进行重构, 生成横截面 pa 图像。PA 成像提供结构和功能信息, 从宏观器官到微观细胞器, 由于存在于体内10的内源性色母象的波长依赖性。PAT 成像已成功地用于乳腺癌检测1, 前哨淋巴结成像11, 氧血红蛋白 (hbo2) 的映射, 脱氧血红蛋白 (HbO), 总血红蛋白浓度 (HbO), 氧饱和度 (so2)12,13、肿瘤血管生成14例, 小动物全身成像15例, 等应用。

Nd:yag/opo 激光器是第一代 PAT 系统的常规激发源, 广泛应用于光声界的小动物成像和深层组织成像16。这些激光器在 ~ 10-100 Hz 的低重复率下提供 ~ 100 mJ 能量脉冲。由于脉冲重复率有限, 使用这些昂贵且笨重的激光的 PAT 成像系统不适合使用单元素超声传感器 (Sut) 进行高速成像。这就抑制了对动物体内高速发生的生理变化的实时监测。使用基于阵列的传感器, 如带有 nd:yag 激光激发的线性、半圆、圆形和体积阵列, 可以进行高速成像。然而, 这些阵列传感器价格昂贵, 与 Sut 相比, 它们的灵敏度较低;然而, 成像速度受到激光低重复率的限制。最先进的单脉冲 PACT 系统与定制的全环阵列传感器获得 PA 数据在 50 Hz 帧速率17.这些阵列传感器需要复杂的后端接收电子和信号放大器, 使整个系统更加昂贵, 临床使用更加困难。

其紧凑的尺寸、较低的成本要求和较高的脉冲重复率 (KHz 顺序) 使脉冲激光二极管 (Pld) 更有前途, 适用于实时成像。由于这些优点, 在第二代 PAT 系统中, Pld 被积极用作备用励磁源。基于pld 的 pat 系统已成功地证明, 高帧率成像使用阵列传感器 18, 深组织和大脑成像19,20,21, 心血管疾病诊断22和风湿病诊断23。由于与阵列传感器相比, Sut 具有高度敏感和更低的成本, 因此它们仍被广泛用于 PAT 成像。基于光纤的 PLD 系统已被证明用于幻象成像 24。以前通过将 PLD 安装在 PAT 扫描仪25内, 已经演示了便携式 pld-pat 系统。使用一个 SUT 圆形扫描仪, 在3次扫描时间内进行幻影成像, 并在5维使用 PLD-PAT 系统19在5个时间内进行大鼠脑成像。

此外, 还对该 pld-pat 系统进行了改进, 使其更加紧凑, 并使用8个基于声学反射的单元素超声波传感器 (subr)26、27创建桌面模型。在这里, Sut 被放置在一个垂直的, 而不是水平的方向, 在90°声学反射器28的帮助下。该系统可用于组织成像和体内小动物大脑成像中的扫描时间高达 0.5 s 和约3厘米深。在这项工作中, 这个桌面 PLD-PAT 系统用于提供实验的视觉演示, 用于在小动物的体内大脑成像和动态可视化的吸收和清除过程的食品和药物管理局 (FDA) 批准的碘苯胺大鼠大脑中的绿色染料 (ICG)。

Protocol

所有动物实验都是根据新加坡南洋理工大学动物护理和使用机构委员会批准的指导方针和条例 (动物议定书编号 ARF-SBS/NIE-0331) 进行的。 1. 系统描述 将 PLD 激光安装到圆形扫描仪中, 并将光学扩散器 (OD) 安装在 PLD 出口窗口的前面, 以使输出光束均匀, 如图 1a 所示。将 PLD 连接到激光驱动器单元 (LDU)。注: PLD 产生 ~ 816 nm 波长脉冲, 脉冲持续时间约 …

Representative Results

该协议展示了所描述的桌面 PLD-PAT 系统在体内动态脑成像中的潜力, 并给出了相应的结果。通过对健康雌性大鼠进行体内脑成像, 证明了桌面 PLD-PAT 系统的高速成像能力。PA 信号是使用 8个 SUTRs 在360°和45°周围的大鼠大脑中旋转收集的, 扫描速度分别为4秒和0.5秒。图 2A. b 显示了一只雌性大鼠 (98 克) 的大脑图像, 扫描速度分别为4秒和0.5秒。在这两个图像中都可?…

Discussion

这项工作提出了一个协议, 使用桌面 PLD-PAT 系统进行实验小动物, 如老鼠体内大脑成像和动态快速吸收和清除过程的造影剂, 如 ICG。笨重、昂贵的 OPO-PAT 系统需要几分钟 (2-5) 才能获得体内单个横截面图像。紧凑、低成本、第一代便携式 PLD-PAT 系统可在5秒内提供单个体内横截面图像。相比之下, 高速、紧凑、低成本的台式机 PLD-PAT 系统仅在 0.5秒26中呈现高质量的2D 横截面体内图像?…

Disclosures

The authors have nothing to disclose.

Acknowledgements

这项研究得到了新加坡卫生部国家医学研究委员会 (NMRC/OFIRGY000/2016:M4062012) 的支持。作者要感谢周伟宏博比先生对机器车间的支持。

Materials

12 V power supply Voltcraft PPS-11810 To supply operating voltage for PLD
Acoustic reflector Olympus F102 45 degree reflector augmented to the ultrasound transducer
Acrylic water tank NTU workshop Custom-made It is used to hold water that acts as an acoustic coupling medium between animal brain and detector
Anesthetic Machine Medical plus pte ltd Non-Rebreathing Anaesthesia machine with oxygen concentrator. Supplies oxygen and isoflurane to animal
Animal distributor In Vivos Pte Ltd, Singapore Animal distributor that supplies small animals for research purpose
Animal holder NTU workshop Custom-made Used for holding animal on its abdomen
Breathing mask NTU workshop Custom-made Used along with animal holder to supply anesthesia mixture to the animal
Circular Scanner NTU workshop Custom-made Scanner is made out of aluminum
DAQ (Data acquisition) Card Spectrum M2i.4932-exp 16 bit, 30 Ms/s, 8 channels, 1 Gs, PCIe
Data acqusition software National Instruments Corporation,Austin,TX,USA) NI LabVIEW 2015 SP1 (32 bit) LabVIEW based program developed in our laboratory for controlling the stepper motor and acquring the PA singnals from the detector
Data processing software Matlab (Mathworks, Natick, MA, USA) Matlab R2015b Matlab code developed in our laboratory for reconstructing cross-sectional PA images
Function generator RIGOL DG1022 To change the repetition rate of the PLD. It will provide TTL signal to synchronize the DAQ with the laser excitation.
Low noise signal amplifier Genetron Custom-made using Mini-circuits, ZFL-500LN-BNC To receive, and amplify the PA signal from SUTR. Its gain is 24 dB.
Optical diffuser Thorlabs DG-120 Used to to make the laser beam homogeneous
Pulsed laser diode Quantel, France QD-Q1924-ILO-WATER It is the excitation laser source with specifications of 816 nm wavelength, 3.4 mJ per pulse energy, 107 ns pulse width, 2 KHz maximum pulse repitition rate, dimensions : 13.0 x 7.6 x 5.0 cm
Rats In Vivos Pte Ltd, Singapore NTac:SD, Sprague Dawley / SD Female, weight 100±10g, strain of rats: Sprague Dawley, age: 4-5 weeks
Stepper motor with gearbox LIN Engineering (Servo Dynamics) Motor: CO-5718L-01P-RO, Gearbox: DPL64/1; Power supply PW-100-24 To move the detector holder in a circular geometry. Torque: 2.08 N-m, Rotor inertia: 2.6 kg-cm2
Ultrasound gel Progress/parker acquasonic gel PA-GEL-CLEA-5000 Clear ultrasound gel
Ultrasound Transducer Olympus V309-SU/ U8423013 Ultrasonic sensors used for photoacoustic detection. Central freqency 5 MHz, 0.5 in
Variable high voltage power supply Elektro-Automatik EA-PS 8160-04 T To change the laser output power
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Tags

Photoacoustic TomographyPATHybrid Biomedical ImagingSmall Animal Brain ImagingPulsed Laser DiodeRat Cortical VasculatureWash-inWash-outDye MonitoringCost-effectivePortableHigh-speed Imaging
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Kalva, S. K., Upputuri, P. K., Rajendran, P., Dienzo, R. A., Pramanik, M. Pulsed Laser Diode-Based Desktop Photoacoustic Tomography for Monitoring Wash-In and Wash-Out of Dye in Rat Cortical Vasculature. J. Vis. Exp. (147), e59764, doi:10.3791/59764 (2019).
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