This work was supported in part by the National Institutes of Health (P41-EB015903, R21EY023043, K25EB015885), National Science of Foundation (CBET-0853773) and Human Frontier Science Program (Young Investigator Grant).

注意:布里渊频谱分析需要单纵模激光（〜10毫瓦的样品）。对于调整的目的，使用激光束（&lt;0.1毫瓦）的强烈衰减部分。 纤维1.初始设置和EMCCD（电子倍增电荷耦合器件）相机<ol><li>识别约1600 mm的自由空间对准了在光学平台上的光谱仪。 </li><li>安装EMCCD相机在自由空间对准的结束。 <ol><li>用螺丝将相机连接到岗位。拧紧后持有人的职位所需的光轴高度。拧紧后持有人使用上表夹在光学平台。 </li></ol></li><li>打开EMCCD相机。要留意避免摄像头饱和。 <ol><li>实验室的计算机上安装摄像头软件，然后将相机连接到电脑。 </li><li>禁用增益设置为低积分时间（〜0.01秒）。开始获取EMCCD图像。观察摄像机图像•计算机屏幕上。 </li></ol></li><li>装入约1600毫米，镜头前的光纤准直器。 <ol><li>放置在光纤准直适配器的光纤准直器。 </li><li>安装适配器到一个职位。拧紧后成柱支架在光轴（摄像机的高度）的近似高度。柱支架拧紧到使用表夹子光学平台。 </li></ol></li><li>对准沿着光轴的输出光束。 <ol><li>使用固定螺钉连接标准虹膜后。拧紧后成柱座。 </li><li>放置虹膜在光纤准直器（&lt;50毫米）的前部。调整光圈的光束的高度。移动沿着期望光轴虹膜，应该直接点到相机。 </li><li>直接的激光输出，如果相机饱和之后，插入一个光密度滤光片。使用光纤准直器的调整螺钉安装到对准沿着光轴的光束。一旦实现这一点，该束将干净地穿过沿整个光路的光圈。 </li></ol></li><li>安装一个匹配的消色差透镜对中（f = 30mm）的在照相机的前面。 </li></ol>光谱仪2.水平阶段<ol><li>挂载水平面具。 <ol><li>使用固定螺钉的水平掩模附着到一个职位。拧紧后成柱座。柱支架安装到一个平移载物台，使掩膜水平滑动进出光束路径。 （图1） </li><li>用有头螺钉拧紧在光学平台上的平移阶段，使得掩模急剧成像到CCD照相机在消色差透镜对的焦点距离（f = 30毫米）。 </li></ol></li><li>安装并对准球面透镜S1（F = 200毫米）600个毫米在水平掩模的前面。 <ol><li>用螺丝安装S1到柱座。拧紧后成柱座。 </li><li>将两片虹膜如1.5.1所述。 placi前纳克S1中的光束路径中，插入前1虹膜和所需的S1位置后的一个光圈（600毫米的水平掩模的前面）。调整虹膜的高度，使得所述光束干净地穿过两者。 </li><li>放置S1 600个毫米的水平掩模的前面（ 图1）。调整高度和S1的角度，直到两个，背反射离开透镜和出未来束，干净地穿过虹膜。拧紧柱支架使用表夹子保持S1到光学平台上。 </li></ol></li><li>安装VIPA2在球面透镜的焦平面（200毫米S1的前面）。 <ol><li> VIPA的支架安装到使用附带的支架螺丝后。拧紧后成柱座。柱支架拧紧到横向平移台。 （图1） </li><li>与入射狭缝垂直定向的VIPA持有人小心地VIPA2。 （图1） </li><li>精细调整位置Of显示VIPA安装到恰好在S1的焦平面。 （通过跟踪束腰带白卡检查。） </li><li>用有头螺钉拧紧平移阶段到光学表和滑动VIPA2用该平移阶段的自由度的光束。 </li></ol></li><li>安装并对准球面透镜S2（F = 200毫米）200毫米的VIPA后和200毫米的水平掩模的前面。按照2.2.1-2.2.3描述的过程。柱支架附着到光学表代替，把它安装到与自由沿光轴取向的其程度的平移阶段。 （图1）使用有头螺钉拧紧平移阶段到光学表。 </li><li>使用水平平移阶段（2.3.4）至VIPA2入口滑入光束路径。观察摄像机图像上的垂直线。 <ol><li>细调整S2使用平移阶段安装在2.4直到在摄像机图像上的垂直线出现尖锐。 </li></ol></li><li>调整与自由对VIPA保持器的水平方向倾斜程度和平移阶段的频谱。使用自由的水平平移度来调整光束的入口位置到标准具。使用自由的水平倾斜程度来调节光束的输入角进标准具。 </li><li>测量技巧和吞吐量。 <ol><li>按F5键或者通过点击在左上角"采集设置"，然后选择该选项，在拍照软件"取图像"拍摄图像。 </li><li>右键点击图片并选择选项"线图"。水平拖动光标在图像生成线图。释放光标来观察产生的线图。 </li><li>使用线图来测量技巧。通过半最大值（FWHM）全宽分隔两个峰之间的距离。瞄准&gt; 30。 </li><li>确定由measurin吞吐量摹与之前和VIPA2后，功率计的功率。目的是为&gt; 50％。 </li><li>调光束的输入角与自由对VIPA支架（2.6）的程度的标准具和观察技巧和吞吐量之间的权衡。 </li></ol></li><li>如果精细度和吞吐量都不能令人满意回去精细调整对齐。确保VIPA2是S1的焦平面。重复步骤2.3.3 -2.7。 </li></ol>光谱仪3.垂直阶段<ol><li>滑动VIPA（VIPA2），排列在第2部分，用该平移阶段（2.3.4）的光束。光谱仪的水平阶段现在将表现为1:1的成像系统。 </li><li>安装垂直面具。 <ol><li>用螺丝将附加的垂直面具到一个职位。拧紧后成直角钳后适配器。拧紧第二次后进入直角适配器，放入柱座。柱支架安装到一个垂直平移阶段，allowi纳克的掩模进出的光束路径的垂直滑动。 （图1） </li><li>使用有头螺钉拧紧垂直平移阶段到光学表，使得垂直方向的荫罩急剧成像到EMCCD照相机200毫米S1的前面。 </li></ol></li><li>安装和对准圆柱透镜C1（F = 200毫米）600个毫米的垂直掩模的前面。 <ol><li>柱面透镜支架螺丝到一个职位。拧紧后成柱座。小心地C1到物镜保持器，并拧紧螺丝固定到位。 </li><li>将两片虹膜如1.5.1所述。前放置C1在光束路径中，插入前1虹膜和所需的S1位置后的一个光圈（600毫米的垂直掩模的前面）。调整虹膜的高度，使得所述光束干净地穿过两者。 </li><li>地方的C1 600个毫米的垂直掩模的前面（ 图1）。仔细调整高度，倾斜，和C1至横向位置两者，背反射离开透镜和出到来的光束，被集中到虹膜。拧紧后持有人持有C1到使用表夹在光学平台。 </li></ol></li><li>安装VIPA1在柱面透镜的焦平面（200毫米的C1的前方）。 <ol><li> VIPA的支架安装到使用附带的支架螺丝后。拧紧后成柱座。柱支架拧紧到垂直平移台。 （图1） </li><li>小心地将VIPA1与入射狭缝水平方向的VIPA持有人。 （图1） </li><li>精细调整VIPA的位置安装以精确地放置VIPA1在C1的焦平面。 （通过跟踪束腰带白卡检查。） </li><li>使用有头螺钉拧紧垂直平移阶段到光学表并滑动VIPA1用该平移阶段的自由度的光束。 </li></ol></li><li>安装和对齐圆柱透镜C2（F = 200毫米）200毫米VIPA1后和200毫米的垂直掩模的前面，以下在3.3.1-3.3.3中描述的方法。柱支架附着到光学表代替，把它安装到与自由沿光轴取向的其程度的平移阶段。 （图1）使用有头螺钉拧紧平移阶段到光学表。 </li><li>使用平移台（3.4.4）至VIPA1入口滑入光束路径。观察摄像机图像上的水平线。 <ol><li>使用平移阶段安装在3.5直到在摄像机图像上的水平线出现尖锐精细调整C2。 </li></ol></li><li>调整与自由对VIPA保持器的垂直倾斜程度和平移阶段的频谱。使用自由的垂直平移度来调整光束的入口位置到标准具。使用自由的垂直方向倾斜程度来调节光束的输入角进等人上。 </li><li>测量技巧和吞吐量。 <ol><li>按照步骤2.7.1 -2.7.2在整个相机屏幕的图像垂直生成线图。 </li><li>使用线路积除以两条水平线之间的距离通过全宽度半最大值（FWHM）来测量手腕。瞄准&gt; 40。 </li><li>通过之前和VIPA1后测量与功率计的功率测量吞吐量。目的是为&gt; 30％。 </li><li>调光束的垂直输入角与自由对VIPA支架（3.7）的垂直方向倾斜程度的标准具。观察技巧和吞吐量之间的权衡。 </li></ol></li><li>如果精细度和吞吐量都不能令人满意回去精细调整对齐。确保VIPA1是C1的焦平面。重复步骤3.4.3-3.8。 </li></ol> 4.组合两个阶段和最后的对的<ol><li>滑动VIPA2。观察水平和垂直方向间隔开的点。这些点的重的单频率激光器的光谱特征。调整东哲行的平移阶段，直到点是大家关注的焦点。 </li><li>测量精细度谱仪和吞吐量。 <ol><li>按照步骤2.7.1.-2.7.2产生斜对面的两个激光点的线图。 </li><li>使用线路积除以两个点之间的距离对角通过全宽度半最大值（FWHM）来测量手腕。瞄准&gt; 30。 </li></ol></li><li>建立一个黑盒子包围光谱仪。 <ol><li>使用施工护栏打造的箱框架，其中应包含整个光谱仪从光纤准直器的CCD摄像机（63英寸×9×15英寸）。 </li><li>覆盖盒子骨架遮光布，然后用胶带紧的角落。确保口罩和东哲行都很方便。 </li></ol></li><li>光纤到一标准的光学探头，连接诸如反射共聚焦显微镜4。标准光学p用于收集背散射光长袍将携带一个布里渊信号。 </li></ol> 5.测量布里渊移<ol><li>接近垂直和水平掩模，直到激光签名消失。相应地移动翻译阶段。 </li><li>使增益和增加相机尽可能的积分时间，而不饱和EMCCD相机。 </li><li>观察样品的布里渊偏移。 <ol><li>放置一个样品中的共焦显微镜（或其它光学探测器）的焦点。的空间分辨率将取决于在共聚焦显微镜中使用的物镜。使用塑料盘或试管液体。使用甲醇为第一测量。 </li></ol></li><li>为了优化分光计吞吐量，打开一个掩模的时间和通过分光计订单通过倾斜VIPA和调整其平移台扫描。发现在该信号出现的最强的顺序。再次关闭屏蔽，直到日Ë激光信号消失。重复与其他掩模和VIPA（在2.6和3.7中描述）。 </li><li>取样品的测量。 <ol><li>取频谱的图像以下步骤2.7.1。 </li><li>通过利用水和玻璃（或与已知的布里渊移其他样品）的光谱的图像获取校准测量。通过单击左上角的"文件"，并选择选项"另存为"保存图像。保存在"的.sif"格式的图像，如果在相机软件进行数据分析。保存图像"的.tif"格式，如果在另一计算软件程序进行数据分析。 </li></ol></li></ol> 6.校准布里渊频谱分析和<ol><li>确定自由光谱范围（FSR）和EMCCD像素中的布里渊光谱仪的光学频率转换比（PR）。 <ol><li>装载数据在相机软件或另一computa其它软件程序。 </li><li>跟随步骤2.7.2以产生跨越所述水校准图像的光谱的线图。 </li><li>适合用洛伦兹曲线对光谱的两个峰，以确定在像素位置（P 水S，P 水-AS）而言峰的位置。另外，手动读取出的峰值位置，采取了最高点的峰值。 </li><li>与玻璃校准图像的频谱重复步骤6.1.2-6.1.3。 </li><li>另外，以6.1.1-6.1.4，同时添加EMCCD框架和适合所有四个峰一次。 （图2） </li><li>用公式1.插入对于p 水AS和P 玻璃AS中6.1.3和6.1.4确定的值计算公关。 Ω 玻璃-AS是已知的29.3 GHz的。在这种情况下，由于自由光谱范围仅为20就会出现混叠为9.3千兆赫的频率偏移千兆赫。为简单起见使用9.3 GHz适用于Ω 玻璃AS。使用7.46 GHz的为Ω水AS。 <img alt="式（1）" class="jove_content" p="" src="/files/ftp_upload/53468/53468eq1.jpg" /></li><li>用公式2.插入对于p 玻璃-S，P 玻璃-AS和 PR，在6.16计算值计算FSR。使用9.3千兆赫的Ω 玻璃AS。 <img alt="公式（2）" class="jove_content" src="/files/ftp_upload/53468/53468eq2.jpg" /></li></ol></li></ol><img alt="图2" class="jove_content" src="/files/ftp_upload/53468/53468fig2.jpg" /> 图2.光谱仪的校准 。从校准样品（A）EMCCD相机帧获得。 （二）洛伦兹曲线拟合（红色）测得的数据（蓝色）。 <a href="https://www.jove.com/files/ftp_upload/53468/53468fig2large.jpg" target="_blank">请点击此处查看该图的放大版本。</a> <ol start="2"><li>确定的布里渊移一个样品。 <ol><li>跟随步骤2.7.2以产生跨越所述样品的光谱行扫描。 </li><li>适合用洛伦兹曲线对光谱的两个峰，以确定在像素位置而言的峰值位置。另外，手动读取出的峰值位置，采取了最高点的峰值。 </li><li>使用下面的等式4和5与先前计算的值，FSR及PR来计算样品的布里渊偏移。 <img alt="公式3" class="jove_content" p="" src="/files/ftp_upload/53468/53468eq3.jpg" /></li></ol></li></ol>

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H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=In+Vivo+Biomechanical+Mapping+of+Normal+and+Keratoconus+Corneas.">In Vivo Biomechanical Mapping of Normal and Keratoconus Corneas.</a> Jama Ophtalmol. , (2015).</li><li>Scarcelli, G., Kim, P., Yun, S. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=+In+Vivo+Measurement+of+Age-Related+Stiffening+in+the+Crystalline+Lens+by+Brillouin+Optical+Microscopy."> In Vivo Measurement of Age-Related Stiffening in the Crystalline Lens by Brillouin Optical Microscopy.</a> Biophys. J. 101 (6), 1539-1545 (2011).</li><li>Antonacci, G., Foreman, M. R., Paterson, C., T&#246;r&#246;k, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Spectral+broadening+in+Brillouin+imaging.">Spectral broadening in Brillouin imaging.</a> Appl. Phys. Lett. 103 (22), 221105 (2013).</li><li>Meng, Z., Traverso, A. J., Yakovlev, V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Background+clean-up+in+Brillouin+microspectroscopy+of+scattering+medium.">Background clean-up in Brillouin microspectroscopy of scattering medium.</a> Opt. Exp. 22 (5), 5410-5415 (2014).</li><li>Reiss, S., Burau, G., Stachs, O., Guthoff, R., Stolz, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Spatially+resolved+Brillouin+spectroscopy+to+determine+the+rheological+properties+of+the+eye+lens.">Spatially resolved Brillouin spectroscopy to determine the rheological properties of the eye lens.</a> Biomed. Opt. Exp. 2 (8), 2144-2159 (2011).</li><li>Scarcelli, G., Kling, S., Quijano, E., Pineda, R., Marcos, S., Yun, S. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Brillouin+microscopy+of+collagen+crosslinking:+noncontact+depth-dependent+analysis+of+corneal+elastic+modulus.">Brillouin microscopy of collagen crosslinking: noncontact depth-dependent analysis of corneal elastic modulus.</a> Invest. Ophthalmol. Vis. Sci. 54 (2), 1418-1425 (2013).</li></ol>

Giuliano Scarcelli and Seok H. Yun hold patents related to Brillouin spectroscopy technology.

这种分光计的配置的一个关键的设计特点是，两个阶段可以独立地对准。当一个VIPA标准具滑出光路，分光计阶段的其余镜片形成1:1的成像系统，使来自每个阶段的频谱图案被成像到CCD照相机。因此，它是直接回到任一个阶段以提高其性能，而不会影响其他级的对准。该组中的协议建议的平移阶段和自由度遵循这一理念，保持独立优化光谱仪的两个阶段的能力。 这是很难装入V...

布里渊散射，首先由Leon布里渊1于1922年描述的，是光从在固体和来自液体或气体的热密度波动的热声模的非弹性散射。的散射光的谱移，通常在子GHz的范围，提供了有关该入射光和与样品中的声子之间的相互作用的信息。其结果是，它可以提供关于被检查材料的粘弹性能的有用信息。 在其自发版本，布里渊散射通常具有横截面在拉曼散射的顺序，产生了非常弱的信号。此外，布里渊频移的幅度比拉曼位移较小的订单。因此，弹性散射光（来自瑞利和米氏散射），杂散光，而反向反射在样本上都可以很容易地掩盖了布里渊光谱特征。故，布里渊光谱仪不仅需要实现的sub-GHz光谱分辨率也高光谱对比度或灭绝。 在传统的布里渊光谱仪这些要求是由扫描光栅单色器，光学打浆方法，以及最普遍地，多遍扫描法布里-珀罗干涉仪2满足。这些方法测量每个频谱分量顺序。这种方法导致了采集时间为单个布里渊频谱范围从几分钟到几个小时，这取决于仪器和在样品上。两阶段VIPA光谱仪，使用该协议建立，具有收集所有频谱分量的内不到一秒，同时提供足够的消光（&gt; 60 dB为单位），以有效地抑制其它杂散信号2的能力。 VIPA的标准具的整合是本光谱仪的关键要素。一个VIPA是一个坚实的标准具有三个不同的C浮动区域:在所述前表面，一个窄抗反射涂层条允许光进入VIPA，而表面的其余部分设有一个高反射（HR）涂层;在后表面的部分反射涂层允许的光的一小部分（〜5％），以进行发送。当聚焦到稍微倾斜VIPA的狭窄的入口时，光束被反射到子组件与VIPA 2内固定的相位差。子组件之间的干扰达到渴望的高光谱色散。在交叉轴配置对两个东哲行依次介绍了正交方向3光谱色散。在正交方向上的光谱色散空间上分离从不需要的串扰，这使得有可能仅拾取布里渊信号的布里渊峰。 图1显示的是两级VIPA分光计的示意图。下面的光学元件中的箭头指示的度自由稀土元素，其中的平移阶段应侧重。 <img alt="图1" class="jove_content" p="" src="/files/ftp_upload/53468/53468fig1.jpg" /> 图1.纯设置。一种光纤开出布里渊散射进入分光光度计。一个柱面透镜C1（F = 200 MM）将光聚焦到第一VIPA（VIPA1）的入口。另一个圆柱透镜的C2（F = 200毫米）映射的光谱角色散成空间分离C2的焦平面。在该平面中，垂直掩模用于选择光谱的所需部分。类似的结构如下，倾斜90度。光束通过球面透镜S1（F = 200毫米）传递并聚焦到第二VIPA（VIPA2）的入口狭缝。甲球面透镜S2（F = 200毫米）产生在其焦平面，其中另一条水平掩模被放置在二维光谱上分离的图案。该HORizontal掩模成像到使用的消色差透镜对的EMCCD相机。 <a href="https://www.jove.com/files/ftp_upload/53468/53468fig1large.jpg" target="_blank">请点击此处查看该图的放大版本。</a> 本科生与一些光学课程和基本取向的经验应该能够建立和使用两个阶段的光谱仪。分光计最近已证明是与各种标准的光学探针的3,4,5（例如，共焦显微镜，内窥镜，裂隙灯检眼镜）兼容。这里，光谱仪被连接到共聚焦显微镜。激光被整合90:10分束器后对准成标准研究倒置显微镜系统。来自样品的背散射光被耦合进单模光纤，使得共焦显微镜。

<table><tbody><tr><td colspan="4">&lt;strong&gt;OPTICS&lt;/strong&gt;</td></tr><tr><td>VIPA (virtual image phase array)</td><td>LIGH MACHINERY</td><td></td><td>Quantity: 2</td></tr><tr><td>Bundle of Three 423 Linear Stages with SM-25 Micrometers</td><td>NEWPORT</td><td>423-MIC&amp;nbsp;</td><td>Quantity: 1</td></tr><tr><td>SS Crossed-Roller Bearing Translation Stage, 0.5 in., 8-32, 1/4-20</td><td>NEWPORT</td><td>9066-X</td><td>Quantity: 1</td></tr><tr><td>Vernier Micrometer, 13&amp;nbsp;mm Travel, 9 lb Load Capacity, 50.8 TPI</td><td>NEWPORT</td><td>SM-13</td><td>Quantity: 1</td></tr><tr><td>Adjustable Width Slit</td><td>NEWPORT</td><td>SV-0.5</td><td>Quantity: 2</td></tr><tr><td>Compact Dovetail Linear Stage, 0.20 in. Z Travel, 1.57x1.57x1.38&amp;nbsp;in.</td><td>NEWPORT</td><td>DS40-Z</td><td>Quantity: 2</td></tr><tr><td>Slotted Base Plate, 25 or 40&amp;nbsp;mm to 65&amp;nbsp;mm Stage, 1.1 in. Range</td><td>NEWPORT</td><td>B-2B</td><td>Quantity: 2</td></tr><tr><td>&amp;Oslash;1/2&quot; Optical Post, 8-32 Setscrew, 1/4&quot;-20 Tap, L = 2&quot;, 5 Pack</td><td>THORLABS</td><td>TR2-P5</td><td>Quantity: 2</td></tr><tr><td>&amp;Oslash;1/2&quot; Post Holders, Spring-Loaded Hex-Locking Thumbscrews, L = 2&quot;, 5 Pack</td><td>THORLABS</td><td>PH2-P5</td><td>Quantity: 1</td></tr><tr><td>&amp;Oslash;1/2&quot; Post Holders, Spring-Loaded Hex-Locking Thumbscrew, L = 3&quot;, 5 Pack</td><td>THORLABS</td><td>PH3-P5</td><td>Quantity: 1</td></tr><tr><td>Imperial Lens Mount For 2&quot; Optics, 8-32 Tap</td><td>THORLABS</td><td>LMR2</td><td>Quantity: 2</td></tr><tr><td>f=200.0 mm, &amp;Oslash;2&quot; Achromatic Doublet, ARC: 400-700 nm</td><td>THORLABS</td><td>AC254-200-A</td><td>Quantity: 2</td></tr><tr><td>Kinematic Mount for up to 1.3&quot; (33&amp;nbsp;mm) Tall Rectangular Optics, Right Handed</td><td>THORLABS</td><td>KM100C</td><td>Quantity: 2</td></tr><tr><td>Fixed Cylindrical Lens Mount, Max Optic Height: 1.60&quot; (40.6 mm)</td><td>THORLABS</td><td>CH1A</td><td>Quantity: 2</td></tr><tr><td>f=200.00 mm, H=30.00 mm, L=32.0 mm, N-BK7 Plano-Convex Cylindrical Lens, Antireflection Coating: 350-700 nm</td><td>THORLABS</td><td>L1653L1-A</td><td>Quantity: 2</td></tr><tr><td>Right-Angle Post Clamp, Fixed 90&amp;deg; Adapter</td><td>THORLABS</td><td>RA90</td><td>Quantity: 1</td></tr><tr><td>Adapter with External C-Mount Threads and Internal SM1 Threads</td><td>THORLABS</td><td>SM1A9</td><td>Quantity: 1</td></tr><tr><td>Studded Pedestal Base Adapter, 1/4&quot;-20 Thread</td><td>THORLABS</td><td>PB4</td><td>Quantity: 2</td></tr><tr><td>Spacer, 2&quot; x 3&quot;, 1.000&quot; Thick</td><td>THORLABS</td><td>Ba2S7</td><td>Quantity: 2</td></tr><tr><td>543 nm, f=15.01 mm, NA=0.17 FC/APC Fiber Collimation Pkg.</td><td>THORLABS</td><td>F260APC-A</td><td>Quantity: 1</td></tr><tr><td>SM1-Threaded Adapter for &amp;Oslash;11&amp;nbsp;mm collimators</td><td>THORLABS</td><td>Ad11F</td><td>Quantity: 1</td></tr><tr><td>Translating Lens Mount for &amp;Oslash;1&quot; Optics, 1 Retaining Ring Included</td><td>THORLABS</td><td>LM1XY</td><td>Quantity: 1</td></tr><tr><td>Single Mode Patch Cable, 450 - 600 nm, FC/APC, 2 m Long</td><td>THORLABS</td><td>P3-460B-FC-2</td><td>Quantity: 1</td></tr><tr><td>1:1 Matched Achr. Pair, f1=30 mm, f2=30 mm, BBAR 400-700 nm</td><td>THORLABS</td><td>MAP103030-A</td><td>Quantity: 1</td></tr><tr><td>SM1 Lens Tube&amp;hellip;length to adjust depend on CCD, we have 3.5&amp;nbsp;inches</td><td>THORLABS</td><td>SM1LXX</td><td>Quantity: 1</td></tr><tr><td>Base Adapters for &amp;Oslash;1/2&quot; Post Holders and &amp;Oslash;1&quot; Posts</td><td>THORLABS</td><td>BE1</td><td>Quantity: 8</td></tr><tr><td>Clamping Forks for&amp;nbsp; &amp;Oslash;1/2&quot; Post Holders and &amp;Oslash;1&quot; Posts</td><td>THORLABS</td><td>CF125</td><td>Quantity: 8</td></tr><tr><td>HW-KIT5 - 4-40 Cap Screw and Hardware Kit for Mini-Series&amp;nbsp;</td><td>THORLABS</td><td>HW-KIT5</td><td>Quantity: 1</td></tr><tr><td>D20S - Standard Iris, &amp;Oslash;20.0 mm Max Aperture&amp;nbsp;</td><td>THORLABS</td><td>D20S</td><td>Quantity: 2</td></tr><tr><td colspan="4">&lt;strong&gt;FOR ENCLOSURE&lt;/strong&gt;</td></tr><tr><td>25 mm Construction Rail, L = 21&quot;</td><td>THORLABS</td><td>XE25L21</td><td>Quantity: 6</td></tr><tr><td>1&quot; Construction Cube with Three 1/4&quot; (M6) Counterbored Holes</td><td>THORLABS</td><td>RM1G</td><td>Quantity: 8</td></tr><tr><td>Right-Angle Bracket for 25 mm Rails</td><td>THORLABS</td><td>XE25A90</td><td>Quantity: 12</td></tr><tr><td>25 mm Construction Rail, L = 15&quot;</td><td>THORLABS</td><td>XE25L15</td><td>Quantity: 4&amp;nbsp;</td></tr><tr><td>25 mm Construction Rail, L = 9&quot;</td><td>THORLABS</td><td>XE25L09</td><td>Quantity: 8</td></tr><tr><td>High Performance Black Masking Tape, 2&quot; x 60 yds. (50 mm x 55 m) Roll</td><td>THORLABS</td><td>T743-2.0</td><td>Quantity: 1</td></tr><tr><td>Low-Profile T-Nut, 1/4&quot;-20 Tapped Hole, Qty: 10</td><td>THORLABS</td><td>XE25T3</td><td>Quantity: 1</td></tr><tr><td>1/4&quot;-20 Low-Profile Channel Screws (100 Screws/Box)</td><td>THORLABS</td><td>SH25LP38</td><td>Quantity: 1</td></tr><tr><td>60&quot; (W) x 3 yds. (L) x 0.005&quot; (T) (1.5 m x 2.7 m x 0.12 mm) Blackout Fabric</td><td>THORLABS</td><td>BK5</td><td>Quantity: 1</td></tr><tr><td colspan="4">&lt;strong&gt;CAMERA, LASER and MICROSCOPE&lt;/strong&gt;</td></tr><tr><td>EMCCD camera</td><td>ANDOR</td><td>iXon Ultra 897</td><td>Quantity: 1</td></tr><tr><td>400 mW single mode green laser</td><td>LASER QUANTUM</td><td>torus 532</td><td>Quantity: 1</td></tr><tr><td>Research Inverted System Microscope&amp;nbsp;</td><td>OLYMPUS</td><td>IX71</td><td>Quantity: 1</td></tr></tbody></table>

high speed sub-ghz spectrometer for brillouin scattering analysis

该协议的目标是建立一个平行的高灭绝和高分辨率光学布里渊光谱仪。布里渊光谱是可以用来获得粘弹性材料属性直接读数的非接触测量方法。它已经在材料特性，结构的监测和环境传感的有用工具。在过去，布里渊光谱已经通常使用扫描法布里 - 珀罗标准具来执行频谱分析。这个过程需要高亮度功率和长采集时间，使得该技术不适合生物医学应用。最近推出的新型光谱仪通过采用两个东哲在交叉轴配置克服了这一难题。这一创新使具有内生物组织的安全界限亚秒采集时间和照明电源子千兆赫（GHz）的分辨率频谱分析。通过这种改进促进了多种新的应用程序立方米rrently正在探索在生物研究和临床应用。

<html> 图3示出有代表性的布里渊频谱和其配合为不同的材料。的东哲行两者的厚度为5毫米，导致大约20千兆赫的FSR。的积分时间这些测量是100毫秒。 100测量是平均。一个校准测量之前获得的光谱进行了拍摄。 <img alt="图3" src="/files/ftp_upload/53468/53468fig3.jpg" /> 图不同材质的3布里渊谱 。 洛伦兹曲线拟合（红?...

Watch this Scientific Journal Video about High Speed Sub-GHz Spectrometer for Brillouin Scattering Analysis at JoVE.com

High Speed Sub-GHz Spectrometer for Brillouin Scattering Analysis

在这里，我们提出了一个协议，以建立一个快速的布里渊光谱仪。级联几乎成像相控阵（VIPA）标准具实现了测量速度比传统扫描法布里 - 珀罗光谱仪快1000倍以上。这种改进提供了用于组织和生物材料在低功率水平的体内布里渊分析。

高速的sub-GHz光谱仪激布里渊散射分析

The goal of this protocol is to build a parallel high-extinction and high-resolution optical Brillouin spectrometer. Brillouin spectroscopy is a ...

high-speed-sub-ghz-spectrometer-for-brillouin-scattering-analysis

Research

JoVE Journal

Bioengineering

14.9K 次观看.  University of Maryland. 在这里，我们提出了一个协议，以建立一个快速的布里渊光谱仪。级联几乎成像相控阵（VIPA）标准具实现了测量速度比传统扫描法布里 - 珀罗光谱仪快1000倍以上。这种改进提供了用于组织和生物材料在低功率水平的体内布里渊分析。 

视频: High Speed Sub-GHz Spectrometer for Brillouin Scattering Analysis

Rejection of Fluorescence Background in Resonance and Spontaneous Raman Microspectroscopy

Raman spectroscopy is often plagued by a strong fluorescent background, particularly for biological samples. If a sample is excited with a train of ultrafast pulses, a system that can temporally separate spectrally overlapping signals on a picosecond timescale can isolate promptly arriving Raman scattered light from late-arriving fluorescence light. Here we discuss the construction and operation of a complex nonlinear optical system that uses all-optical switching in the form of a low-power optical Kerr gate to isolate Raman and fluorescence signals. A single 808 nm laser with 2.4 W of average power and 80 MHz repetition rate is split, with approximately 200 mW of 808 nm light being converted to &lt; 5 mW of 404 nm light sent to the sample to excite Raman scattering. The remaining unconverted 808 nm light is then sent to a nonlinear medium where it acts as the pump for the all-optical shutter. The shutter opens and closes in 800 fs with a peak efficiency of approximately 5%. Using this system we are able to successfully separate Raman and fluorescence signals at an 80 MHz repetition rate using pulse energies and average powers that remain biologically safe. Because the system has no spare capacity in terms of optical power, we detail several design and alignment considerations that aid in maximizing the throughput of the system. We also discuss our protocol for obtaining the spatial and temporal overlap of the signal and pump beams within the Kerr medium, as well as a detailed protocol for spectral acquisition. Finally, we report a few representative results of Raman spectra obtained in the presence of strong fluorescence using our time-gating system.

We discuss the construction and operation of a complex nonlinear optical
system that uses ultrafast all-optical switching to isolate Raman from fluorescence
signals. Using this system we are able to successfully separate Raman and fluorescence
signals utilizing pulse energies and average powers that remain biologically safe.

波谱与自发拉曼显微荧光背景的抑制

Raman spectroscopy is often plagued by a strong fluorescent background, particularly for biological samples. If a sample is excited with a train of ...

科研

生物工程

Fabrication And Characterization Of Photonic Crystal Slow Light Waveguides And Cavities

Slow light has been one of the hot topics in the photonics community in the past decade, generating great interest both from a fundamental point of view and for its considerable potential for practical applications. Slow light photonic crystal waveguides, in particular, have played a major part and have been successfully employed for delaying optical signals1-4 and the enhancement of both linear5-7 and nonlinear devices.8-11
Photonic crystal cavities achieve similar effects to that of slow light waveguides, but over a reduced band-width. These cavities offer high Q-factor/volume ratio, for the realization of optically12 and electrically13 pumped ultra-low threshold lasers and the enhancement of nonlinear effects.14-16 Furthermore, passive filters17 and modulators18-19 have been demonstrated, exhibiting ultra-narrow line-width, high free-spectral range and record values of low energy consumption.
To attain these exciting results, a robust repeatable fabrication protocol must be developed. In this paper we take an in-depth look at our fabrication protocol which employs electron-beam lithography for the definition of photonic crystal patterns and uses wet and dry etching techniques. Our optimised fabrication recipe results in photonic crystals that do not suffer from vertical asymmetry and exhibit very good edge-wall roughness. We discuss the results of varying the etching parameters and the detrimental effects that they can have on a device, leading to a diagnostic route that can be taken to identify and eliminate similar issues.
The key to evaluating slow light waveguides is the passive characterization of transmission and group index spectra. Various methods have been reported, most notably resolving the Fabry-Perot fringes of the transmission spectrum20-21 and interferometric techniques.22-25 Here, we describe a direct, broadband measurement technique combining spectral interferometry with Fourier transform analysis.26 Our method stands out for its simplicity and power, as we can characterise a bare photonic crystal with access waveguides, without need for on-chip interference components, and the setup only consists of a Mach-Zehnder interferometer, with no need for moving parts and delay scans.
When characterising photonic crystal cavities, techniques involving internal sources21 or external waveguides directly coupled to the cavity27 impact on the performance of the cavity itself, thereby distorting the measurement. Here, we describe a novel and non-intrusive technique that makes use of a cross-polarised probe beam and is known as resonant scattering (RS), where the probe is coupled out-of plane into the cavity through an objective. The technique was first demonstrated by McCutcheon et al.28 and further developed by Galli et al.29

Use of photonic crystal slow light waveguides and cavities has been widely adopted by the photonics community in many differing applications. Therefore fabrication and characterization of these devices are of great interest. This paper outlines our fabrication technique and two optical characterization methods, namely: interferometric (waveguides) and resonant scattering (cavities).

光子晶体慢光波导和腔体的制备与表征

Slow light has been one of the hot topics in the photonics community in the past decade, generating great interest both from a fundamental point of view ...

工程学

Measurement of Scattering Nonlinearities from a Single Plasmonic Nanoparticle

Plasmonics, which are based on the collective oscillation of electrons due to light excitation, involve strongly enhanced local electric fields and thus have potential applications in nonlinear optics, which requires extraordinary optical intensity. One of the most studied nonlinearities in plasmonics is nonlinear absorption, including saturation and reverse saturation behaviors. Although scattering and absorption in nanoparticles are closely correlated by the Mie theory, there has been no report of nonlinearities in plasmonic scattering until very recently. 
Last year, not only saturation, but also reverse saturation of scattering in an isolated plasmonic particle was demonstrated for the first time. The results showed that saturable scattering exhibits clear wavelength dependence, which seems to be directly linked to the localized surface plasmon resonance (LSPR). Combined with the intensity-dependent measurements, the results suggest the possibility of a common mechanism underlying the nonlinear behaviors of scattering and absorption. These nonlinearities of scattering from a single gold nanosphere (GNS) are widely applicable, including in super-resolution microscopy and optical switches. 
In this paper, it is described in detail how to measure nonlinearity of scattering in a single GNP and how to employ the super-resolution technique to enhance the optical imaging resolution based on saturable scattering. This discovery features the first super-resolution microscopy based on nonlinear scattering, which is a novel non-bleaching contrast method that can achieve a resolution as low as l/8 and will potentially be useful in biomedicine and material studies.

Saturable and reverse saturable scattering were discovered in isolated plasmonic particles and adopted as a novel non-bleaching contrast method in super-resolution microscopy. Here the experimental procedures of detecting and extracting nonlinear scattering are explained in detail, as well as how to enhance resolution with the aid of saturated excitation microscopy.

从单电浆纳米粒子散射非线性测量

Plasmonics, which are based on the collective oscillation of electrons due to light excitation, involve strongly enhanced local electric fields and thus ...

High Resolution Phonon-assisted Quasi-resonance Fluorescence Spectroscopy

High resolution optical spectroscopy methods are demanding in terms of either technology, equipment, complexity, time or a combination of these. Here we demonstrate an optical spectroscopy method that is capable of resolving spectral features beyond that of the spin fine structure and homogeneous linewidth of single quantum dots (QDs) using a standard, easy-to-use spectrometer setup. This method incorporates both laser and photoluminescence spectroscopy, combining the advantage of laser line-width limited resolution with multi-channel photoluminescence detection. Such a scheme allows for considerable improvement of resolution over that of a common single-stage spectrometer. The method uses phonons to assist in the measurement of the photoluminescence of a single quantum dot after resonant excitation of its ground state transition. The phonon&#39;s energy difference allows one to separate and filter out the laser light exciting the quantum dot. An advantageous feature of this method is its straight forward integration into standard spectroscopy setups, which are accessible to most researchers.

The manuscript describes a method of phonon-assisted quasi-resonant fluorescence spectroscopy that incorporates both laser-limited resolution and photoluminescence (PL) spectroscopy. This method utilizes optical phonons to provide linewidth-limited resolution spectra of atom-like semiconductor structures in the energy domain. The method is also easily realized with a single spectrometer optical spectroscopy setup.

高分辨率声子辅助准共振荧光光谱仪

High resolution optical spectroscopy methods are demanding in terms of either technology, equipment, complexity, time or a combination of these. Here we ...

Non-equilibrium Microwave Plasma for Efficient High Temperature Chemistry

A flowing microwave plasma based methodology for converting electric energy into internal and/or translational modes of stable molecules with the purpose of efficiently driving non-equilibrium chemistry is discussed. The advantage of a flowing plasma reactor is that continuous chemical processes can be driven with the flexibility of startup times in the seconds timescale. The plasma approach is generically suitable for conversion/activation of stable molecules such as CO2, N2 and CH4. Here the reduction of CO2 to CO is used as a model system: the complementary diagnostics illustrate how a baseline thermodynamic equilibrium conversion can be exceeded by the intrinsic non-equilibrium from high vibrational excitation. Laser (Rayleigh) scattering is used to measure the reactor temperature and Fourier Transform Infrared Spectroscopy (FTIR) to characterize in situ internal (vibrational) excitation as well as the effluent composition to monitor conversion and selectivity.

This article describes a flowing microwave reactor that is used to drive efficient non-equilibrium chemistry for the application of conversion/activation of stable molecules such as CO2, N2 and CH4. The goal of the procedure described here is to measure the in situ gas temperature and gas conversion.

用于高效高温化学的非平衡微波等离子体

A flowing microwave plasma based methodology for converting electric energy into internal and/or translational modes of stable molecules with the purpose ...

化学

High-speed Continuous-wave Stimulated Brillouin Scattering Spectrometer for Material Analysis

Recent years have witnessed a significant increase in the use of spontaneous Brillouin spectrometers for non-contact analysis of soft matter, such as aqueous solutions and biomaterials, with fast acquisition times. Here, we discuss the assembly and operation of a Brillouin spectrometer that uses stimulated Brillouin scattering (SBS) to measure stimulated Brillouin gain (SBG) spectra of water and lipid emulsion-based tissue-like samples in transmission mode with &lt;10 MHz spectral-resolution and &lt;35 MHz Brillouin-shift measurement precision at &lt;100 ms. The spectrometer consists of two nearly counter-propagating continuous-wave (CW) narrow-linewidth lasers at 780 nm whose frequency detuning is scanned through the material Brillouin shift. By using an ultra-narrowband hot rubidium-85 vapor notch filter and a phase-sensitive detector, the signal-to-noise-ratio of the SBG signal is significantly enhanced compared to that obtained with existing CW-SBS spectrometers. This improvement enables measurement of SBG spectra with up to 100-fold faster acquisition times, thereby facilitating high spectral-resolution and high-precision Brillouin analysis of soft materials at high speed.

We describe the construction of a rapid continuous-wave-stimulated-Brillouin-scattering (CW-SBS) spectrometer. The spectrometer employs single-frequency diode-lasers and an atomic vapor notch-filter to acquire transmission spectra of turbid/non-turbid samples with high spectral-resolution at speeds up to 100-fold faster than those of existing CW-SBS spectrometers. This improvement enables high-speed Brillouin material analysis.

用于材料分析的高速连续波受激布里渊散射光谱仪

Recent years have witnessed a significant increase in the use of spontaneous Brillouin spectrometers for non-contact analysis of soft matter, such as ...

Observation and Analysis of Blinking Surface-enhanced Raman Scattering

From a single molecule at a silver nanoaggregate junction, blinking surface-enhanced Raman scattering (SERS) is observed. Here, a protocol is presented on how to prepare the SERS-active silver nanoaggregate, record a video of certain blinking spots in the microscopic image, and analyze the blinking statistics. In this analysis, a power law reproduces the probability distributions for bright events relative to their duration. The probability distributions for dark events are fitted by a power law with an exponential function. The parameters of the power law represent molecular behavior in both bright and dark states. The random walk model and the speed of the molecule across the entire silver surface can be estimated. It is difficult to estimate even when using averages, autocorrelation functions, and super-resolution SERS imaging. In the future, power law analyses should be combined with spectral imaging, because the origins of blinking cannot be confirmed by this analysis method alone.

This protocol describes the analysis of blinking surface-enhanced Raman scattering due to the random walk of a single molecule on a silver surface using power laws.

闪烁表面增强拉曼散射的观测与分析

From a single molecule at a silver nanoaggregate junction, blinking surface-enhanced Raman scattering (SERS) is observed. Here, a protocol is presented on ...

Quantitative Analysis of Vacuum Induction Melting by Laser-induced Breakdown Spectroscopy

Vacuum induction melting is a popular method for refining high purity metal and alloys. Traditionally, standard process control in metallurgy involves several steps, include drawing samples, cooling, cutting, transport to the laboratory, and analysis. The whole analysis process requires more than 30 minutes, which hinders on-line process control. Laser-induced breakdown spectroscopy is an excellent on-line analysis method that can satisfy the requirements of vacuum induction melting because it is fast and noncontact and does not require sample preparation. The experimental facility uses a lamp-pumped Q-switched laser to ablate melted liquid steel with an output energy of 80 mJ, a frequency of 5 Hz, a FWHM pulse width of 20 ns, and a working wavelength of 1,064 nm. A multi-channel linear charge coupled device (CCD) spectrometer is used to measure the emission spectrum in real time, with a spectral range from 190 to 600 nm and a resolution of 0.06 nm at a wavelength of 200 nm. The protocol includes several steps: standard alloy sample preparation and an ingredient test, smelting of standard samples and determination of the laser breakdown spectrum, and construction of the elements concentration quantitative analysis curve of each element. To realize the concentration analysis of unknown samples, the spectrum of a sample also needs to be measured and disposed with the same process. The composition of all main elements in the melted alloy can be quantitatively analyzed with an internal standard method. The calibration curve shows that the limit of detection of most metal elements ranges from 20-250 ppm. The concentration of elements, such as Ti, Mo, Nb, V, and Cu, can be lower than 100 ppm, and the concentrations of Cr, Al, Co, Fe, Mn, C, and Si range from 100-200 ppm. The R2 of some calibration curves can exceed 0.94.

During vacuum induction melting, laser-induced breakdown spectroscopy is used to perform real-time quantitative analysis of the main-ingredient elements of a molten alloy.

激光击穿光谱对真空感应熔融的定量分析

Vacuum induction melting is a popular method for refining high purity metal and alloys. Traditionally, standard process control in metallurgy involves ...

Optimizing Brillouin Optical Time-Domain Analyzers Based on Gain Spectrum Engineering

Demonstrated is a unique method for sensing performance enhancement in Brillouin optical time-domain analyzers (BOTDA). A Brillouin gain spectrum (BGS) is superimposed with two symmetric Brillouin loss spectra (BLS). This leads to a complex engineered spectrum shape that is more resistant to the sensing system noise. Instead of only one pump and probe interaction as in the conventional BOTDA setup, three optical probe waves are exploited, with one probe located in the BGS and the other two symmetrically in the BLS. Due to the resistance and insensitivity of the engineered spectrum shape to the noise, the sensing performance is enhanced by 60% and the measurand resolution is doubled.

A protocol for Brillouin optical time-domain analyzers based on gain spectrum engineering is presented. Enhancements in the sensing performance, including sensing range and measurand resolution are achieved and the excess Brillouin intensity noise is studied. The protocol introduces a new way to enhance distributed Brillouin sensing performance.

Demonstrated is a unique method for sensing performance enhancement in Brillouin optical time-domain analyzers (BOTDA). A Brillouin gain spectrum (BGS) is ...

Preparation of Extracellular Matrix Protein Fibers for Brillouin Spectroscopy

Brillouin spectroscopy is an emerging technique in the biomedical field. It probes the mechanical properties of a sample through the interaction of visible light with thermally induced acoustic waves or phonons propagating at a speed of a few km/sec. Information on the elasticity and structure of the material is obtained in a nondestructive contactless manner, hence opening the way to in vivo applications and potential diagnosis of pathology. This work describes the application of Brillouin spectroscopy to the study of biomechanics in elastin and trypsin-digested type I collagen fibers of the extracellular matrix. Fibrous proteins of the extracellular matrix are the building blocks of biological tissues and investigating their mechanical and physical behavior is key to establishing structure-function relationships in normal tissues and the changes which occur in disease. The procedures of sample preparation followed by measurement of Brillouin spectra using a reflective substrate are presented together with details of the optical system and methods of spectral data analysis.

We present a protocol for the application of Brillouin light scattering spectroscopy to elastin and trypsin-purified type I collagen fibers of the extracellular matrix to extract their full elastic properties.

细胞外基质蛋白纤维的布里渊谱的制备

Brillouin spectroscopy is an emerging technique in the biomedical field. It probes the mechanical properties of a sample through the interaction of ...