Name: using an automated hirschberg test app to evaluate ocular alignment
Uploaded: 2020-03-24T15:00:00
Description: Watch this Scientific Journal Video about Using an Automated Hirschberg Test App to Evaluate Ocular Alignment at JoVE.com

这项工作得到了NIH授予R44EY025902和大众眼耳固化儿童补助金部分支持。

这项研究是根据《赫尔辛基宣言》的原则，在舍彭斯眼科研究所（马萨诸塞州波士顿）和斯波尔丁康复医院（马萨诸塞州波士顿）进行的。获得所有参与者的知情同意。这项研究得到了当地质量眼耳机构审查委员会（马萨诸塞州波士顿）的批准。
注：患者纳入标准是水平斜体（恒定或间歇性外向或致向）的先前诊断，没有其他视力障碍。这项研究是先前12日...

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没有专业培训的人可以使用 EyeTurn 应用程序来捕捉眼睛的图片并获得眼部对准测量，现场或远程的眼部护理专家可能会对此进行解释。该应用程序仅提供失调的大小，而不是任何解释或诊断。眼科专家（如视光师或眼科医生）应确定失调是否显著，并考虑其他因素（包括测量条件）后做出诊断。
拍摄高质量的照片对于测量至关重要。相机应放置在两只眼睛之间的位置。离中?...

在视力护理诊所经常进行眼部对准测量。棱镜中和覆盖测试是量化眼睛失调程度（斜视）的常用临床方法。这种方法需要高度的训练和经验。当患者不能完全参与检查时，如幼儿1、脑损伤或中风2或发育障碍3，准确的测量就更具挑战性。此外，在学校筛查中需要进行眼对对齐测试，因为在儿童时期，美国4人口中约有5-8%的人患有斜视，是弱视的一个巨大危险因素，大约30-40%的弱视病例归因于斜视5、6、7。6,75然而，学校护士通常没有受过培训，以进行标准覆盖测试与棱镜中和这种筛选。对于非眼部护理专业人员来说，斜视筛查的另一个挑战是间歇性斜视（不均匀并不总是表现出）和较小的错位幅度在视觉上并不明显（<15 棱镜屈光剂 []）8。
为了解决斜视在检测和测量方面的挑战，我们开发了一款智能手机应用程序（EyeTurn），通过比较眼睛之间角膜反射的位移来实现和自动化摄影Hirschberg方法9。虽然传统的摄影Hirschberg方法已被证明在诊所10，11,11具有良好的可重复性，但专用的独立设备的成本是广泛采用的障碍。通过提供一个易于使用的工具来测量眼睛与标准智能手机的对齐，我们假设它将被广泛采用在学校视力筛查和非眼科护理专业人员使用。我们以前的评估研究表明，应用测量与目前棱镜的临床标准和备用覆盖测试12一致，对于高达60Μ的分裂性致向和外向性。在一项试点的学校筛查研究中，我们还显示，该应用程序可以帮助学校护士检测间歇性外生炎的儿童，他们错过了标准学校视力筛查协议13。
出于研究目的，研究人员和临床医生目前可应要求提供该应用程序的 iOS 版本。到目前为止，请求者包括学校护士、儿科眼科医生、视光师、神经眼科医生和斜视专家。本文的目的是共享详细的应用协议，用于使用应用程序评估不同查看条件下的眼部对齐，即近距和远固定距离;带和没有眼罩打破双目融合。

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>EyeTurn</td>
			<td>EyeNexo</td>
			<td></td>
			<td>Smartphone app for measureing eye misalignment</td>
		</tr>
	</tbody>
</table>

using an automated hirschberg test app to evaluate ocular alignment

已开发一个智能手机应用程序，用于执行自动摄影 Hirschberg 测试，以客观测量眼部失调。通过计算手机摄像头闪光灯相对于光圈中心产生的角膜反射相对于高分辨率图像的差异，该应用程序可以测量失调，其精度远高于执行 Hirschberg 测试的肉眼。与临床金标准棱镜和替代盖试验相比，在以前的临床评价研究中得到了验证。本文的目的是描述有关如何使用应用程序测量不同固定距离的眼部对齐，没有或用封面来打破融合，以及角度 kappa 的测试技术，以便用户可以使用该应用程序执行等效测试通常在诊所使用棱镜进行。

在本作品中，我们描述了使用执行摄影 Hirschberg 测试的智能手机应用程序评估眼部对准的协议。应用程序的界面如图1所示。用户可以选择执行盖测试或测量患者，两只眼睛同时固定在目标，无论是接近或远的固定距离。根据测试目的确定查看条件后，用户可以按照协议并拍摄患者的照片。图像处理后，应用程序将向用户显示分析结果。如图<strong clas...

Watch this Scientific Journal Video about Using an Automated Hirschberg Test App to Evaluate Ocular Alignment at JoVE.com

Using an Automated Hirschberg Test App to Evaluate Ocular Alignment

我们提出了使用智能手机应用程序执行 Hirschberg 测试的协议，用于在近和远固定条件下测量骨骼和间歇性眼部错位（斜体）。

使用自动赫希伯格测试应用程序评估眼部对齐

A smartphone app has been developed to perform the automated photographic Hirschberg test for objective measurement of ocular misalignment. By computing ...

我们开发的移动应用程序可用于测量用于研究或临床用途的斜视。这种方法，便于轻松的斜视测量，目前需要大量的培训和经验，仅使用标准手机摄像头和应用程序软件执行。在应用界面中，用户可以选择执行覆盖测试，或同时测量双眼固定在目标上的患者，距离接近或远。要测量视点，使用单个快照，并近乎固定，请启动应用程序，将模式设置为"无遮盖"。选择"靠近"，将手机定位在距离患者约 40 厘米的横向方向，后置摄像头面向患者。指示患者将手电筒固定在手电筒上，此时手电筒应关闭。对于需要精确住宿的测试，请将固定目标粘贴在手机的背面，直接在手电筒的正下方或上方。当患者确认他们正在修复时，按圆形按钮拍摄要由应用分析的快照。分析完成后，应用程序将显示检测到的肢体边界、眼睛的中心和角膜反射的位置。在验证这些特征是否检测到无明显错误后，观察与眼睛对齐相关的测量结果。然后，按"保存"按钮将当前测试保存在手机中，或按后退箭头按钮重新测试。要测量应用中远固定时间歇性眼部不对齐，请将模式设置为"覆盖测试"并选择"远"。将手机保持横向，距离患者眼睛水平约 40 厘米，闪光灯位于患者眼睛之间。指示患者在电话上方查看大约六米外的目标。让患者遮住一只眼睛，按下圆形按钮，应用程序将开始检测。在确保患者正确固定的同时，请让他们快速消除遮挡。一旦消除遮挡，应用程序将自动拍照。分析完成后，应用程序将显示检测到的肢体边界、眼睛的中心和角膜反射的位置。在验证未发现这些特征时没有明显的错误后，在保存数据或重复测试之前，请查看棱镜二点中眼睛对齐的测量结果。对于应用中的"角卡帕"测量，请选择测量卡帕角度。将手机定位在距离患者约 40 厘米的景观方向，并指示患者使用眼睛进行测试，以固定手电筒，同时覆盖另一只眼睛。在确保患者正确固定时，点按圆形按钮拍摄快照，该快照将由应用进行分析。分析完成后，应用程序将显示检测到的肢体边界、眼睛的中心和角膜反射的位置。在验证未检测到这些特征时没有明显的错误后，请查看 Kappa 角度的测量结果（度）。在此代表性分析中，正确检测了两只眼睛的肢体边界和手电筒的角膜反射，表明眼对准测量不受图像分析误差影响。在这个特殊的情况下，患者已经离开了外向性，这从左眼较大的角膜反射偏移中可以明显看出。应用程序没有报告哪个眼睛偏离了，但是，因为在一个小的斜视角度和未知的Kappa角度的情况下，这将是不可靠的应用程序来确定偏离的眼睛。为了进行比较，这里显示了一个没有斜视的示例。在此分析中，可以观察到错误的肢体检测。虽然角膜反射的检测正确，但绿色圆圈显然与边缘边界不匹配，表示应重复测试。在这项对14个受试者的分析中，10个受试者表现出外向性，4个受试者表现出外向性。线性回归分析，显示眼部斜角的测量，与临床覆盖测试测量一致。使用该应用程序时，需要记住两件重要的事情。一、正确设置每个特定测试的盖板测试和固定模式。二、检查眼部功能检测是否正确。该应用程序只允许斜视测量，它不给诊断。临床医生需要将眼部测量与其他视力测量相结合，以做出诊断。

using-an-automated-hirschberg-test-app-to-evaluate-ocular-alignment

Research

JoVE Journal

Medicine

An in vivo Assay to Test Blood Vessel Permeability

This method is based on the intravenous injection of Evans Blue in mice as the test animal model. Evans blue is a dye that binds albumin. Under physiologic conditions the endothelium is impermeable to albumin, so Evans blue bound albumin remains restricted within blood vessels. In pathologic conditions that promote increased vascular permeability endothelial cells partially lose their close contacts and the endothelium becomes permeable to small proteins such as albumin. This condition allows for extravasation of Evans Blue in tissues. A healthy endothelium prevents extravasation of the dye in the neighboring vascularized tissues. Organs with increased permeability will show significantly increased blue coloration compared to organs with intact endothelium. The level of vascular permeability can be assessed by simple visualization or by quantitative measurement of the dye incorporated per milligram of tissue of control versus experimental animal/tissue. Two powerful aspects of this assay are its simplicity and quantitative characteristics. Evans Blue dye can be extracted from tissues by incubating a specific amount of tissue in formamide. Evans Blue absorbance maximum is at 620 nm and absorbance minimum is at 740 nm. By using a standard curve for Evans Blue, optical density measurements can be converted into milligram dye captured per milligram of tissue. Statistical analysis should be used to assess significant differences in vascular permeability.

An in vivo Assay to Test Blood Vessel Permeability

We are presenting an in vivo assay to test blood vessel permeability. This assay is based on intravenous injection of a dye and subsequent visualization of its diffusion into interstitial spaces.

一个<em在体内</em分析测试血管通透性

This method is based on the intravenous injection of Evans Blue in mice as the test animal model. Evans blue is a dye that binds albumin. Under ...

科研

医学

Using Quantitative Real-time PCR to Determine Donor Cell Engraftment in a Competitive Murine Bone Marrow Transplantation Model

Murine bone marrow transplantation models provide an important tool in measuring hematopoietic stem cell (HSC) functions and determining genes/molecules that regulate HSCs. In these transplant model systems, the function of HSCs is determined by the ability of these cells to engraft and reconstitute lethally irradiated recipient mice. Commonly, the donor cell contribution/engraftment is measured by antibodies to donor- specific cell surface proteins using flow cytometry. However, this method heavily depends on the specificity and the ability of the cell surface marker to differentiate donor-derived cells from recipient-originated cells, which may not be available for all mouse strains. Considering the various backgrounds of genetically modified mouse strains in the market, this cell surface/ flow cytometry-based method has significant limitations especially in mouse strains that lack well-defined surface markers to separate donor cells from congenic recipient cells. Here, we reported a PCR-based technique to determine donor cell engraftment/contribution in transplant recipient mice. We transplanted male donor bone marrow HSCs to lethally irradiated congenic female mice. Peripheral blood samples were collected at different time points post transplantation. Bone marrow samples were obtained at the end of the experiments. Genomic DNA was isolated and the Y chromosome specific gene, Zfy1, was amplified using quantitative Real time PCR. The engraftment of male donor-derived cells in the female recipient mice was calculated against standard curve with known percentage of male vs. female DNAs. Bcl2 was used as a reference gene to normalize the total DNA amount. Our data suggested that this approach reliably determines donor cell engraftment and provides a useful, yet simple method in measuring hematopoietic cell reconstitution in murine bone marrow transplantation models. Our method can be routinely performed in most laboratories because no costly equipment such as flow cytometry is required.

Determining donor cell engraftment presents a challenge in mouse bone marrow transplant models that lack well-defined phenotypical markers. We described a methodology to quantify male donor cell engraftment in female transplant recipient mice. This method can be used in all mouse strains for the study of HSC functions.

采用实时定量PCR检测供体细胞植入一个具有竞争力的小鼠骨髓移植模型

Murine bone marrow transplantation models provide an important tool in measuring hematopoietic stem cell (HSC) functions and determining genes/molecules ...

Using a Knee Arthrometer to Evaluate Tissue-specific Contributions to Knee Flexion Contracture in the Rat

Normal knee range of motion (ROM) is critical to well-being and allows one to perform basic activities such as walking, climbing stairs and sitting. Lost ROM is called a joint contracture and results in increased morbidity. Due to the difficulty of reversing established knee contractures, early detection is important, and hence, knowing risk factors for their development is essential. The rat represents a good model with which the effect of an intervention can be studied due to the similarity of rat knee anatomy to that of humans, the rat's ability to tolerate long durations of knee immobilization in flexion, and because mechanical data can be correlated with histologic and biochemical analysis of knee tissue.
Using an automated arthrometer, we demonstrate a validated, precise, reproducible, user-independent method of measuring the extension ROM of the rat knee joint at specific torques. This arthrometer can be used to determine the effects of interventions on knee joint ROM in the rat.

The goal of the protocol is to measure the extension range of motion of the rat knee. The effects of various diseases that increase the stiffness of the knee joint and the effectiveness of treatments can be quantified.

用膝关节关节仪评价大鼠膝关节屈曲收缩的特定组织贡献

Normal knee range of motion (ROM) is critical to well-being and allows one to perform basic activities such as walking, climbing stairs and sitting. Lost ...

Sucrose Preference and Novelty-Induced Hypophagia Tests in Rats using an Automated Food Intake Monitoring System

The prevalence and incidence of depressive disorders are rising worldwide, affecting about 322 million individuals, underlining the need for behavioral studies in animal models. In this protocol, to study depression-like and anhedonic behavior in rats, the established sucrose preference and novelty-induced hypophagia tests are combined with an automated food and liquid intake monitoring system. Prior to testing, in the sucrose preference paradigm, male rats are trained for at least 2 days to consume a sucrose solution in addition to tap water. During the test, rats are again exposed to water and sucrose solution. Consumption is registered every second by the automated system. The ratio of sucrose to total water intake (sucrose preference ratio) is a surrogate parameter for anhedonia. In the novelty-induced hypophagia test, male rats undergo a training period in which they are exposed to a palatable snack. During training, rodents show a stable baseline snack intake. On test day, the animals are transferred from home cages into a fresh, empty cage representing a novel unknown environment with access to the known palatable snack. The automated system records the total intake and its underlying microstructure (e.g., latency to approaching the snack), providing insight into anhedonic and anxious behaviors. The combination of these paradigms with an automated measuring system provides more detailed information, along with higher accuracy by reducing measuring errors. However, the tests use surrogate parameters and only depict depression and anhedonia in an indirect manner.

Presented here is a protocol to study depression-like and anhedonic behavior in rats. It combines two well-established behavioral methods, the sucrose preference and novelty-induced hypophagia tests, with an automated food and liquid intake monitoring system, to indirectly investigate rodent behavior using surrogate parameters.

使用自动食物摄入量监测系统对大鼠的蔗糖偏好和新奇诱导性低吞咽测试

The prevalence and incidence of depressive disorders are rising worldwide, affecting about 322 million individuals, underlining the need for behavioral ...

Endothelial Cell Transcytosis Assay as an In Vitro Model to Evaluate Inner Blood-Retinal Barrier Permeability

Dysfunction of the blood-retinal barrier (BRB) contributes to the pathophysiology of several vascular eye diseases, often resulting in retinal edema and subsequent vision loss. The inner blood-retinal barrier (iBRB) is mainly composed of retinal vascular endothelium with low permeability under physiological conditions. This feature of low permeability is tightly regulated and maintained by low rates of paracellular transport between adjacent retinal microvascular endothelial cells, as well as transcellular transport (transcytosis) through them. The assessment of retinal transcellular barrier permeability may provide fundamental insights into iBRB integrity in health and disease. In this study, we describe an endothelial cell (EC) transcytosis assay, as an in vitro model for evaluating iBRB permeability, using human retinal microvascular endothelial cells (HRMECs). This assay assesses the ability of HRMECs to transport transferrin and horseradish peroxidase (HRP) in receptor- and caveolae-mediated transcellular transport processes, respectively. Fully confluent HRMECs cultured on porous membrane were incubated with fluorescent-tagged transferrin (clathrin-dependent transcytosis) or HRP (caveolae-mediated transcytosis) to measure the levels of transferrin or HRP transferred to the bottom chamber, indicative of transcytosis levels across the EC monolayer. Wnt signaling, a known pathway regulating iBRB, was modulated to demonstrate the caveolae-mediated HRP-based transcytosis assay method. The EC transcytosis assay described here may provide a useful tool for investigating the molecular regulators of EC permeability and iBRB integrity in vascular pathologies and for screening drug delivery systems.

Endothelial Cell Transcytosis Assay as an In Vitro Model to Evaluate Inner Blood-Retinal Barrier Permeability

This protocol illustrates an in vitro endothelial cell transcytosis assay as a model to evaluate inner blood-retinal barrier permeability by measuring the ability of human retinal microvascular endothelial cells to transport horseradish peroxidase across cells in caveolae-mediated transcellular transport processes.

内皮细胞转吞测定作为 体外 模型评估内部血-视网膜屏障通透性

Dysfunction of the blood-retinal barrier (BRB) contributes to the pathophysiology of several vascular eye diseases, often resulting in retinal edema and ...

Semi-Automated Planimetric Quantification of Dental Plaque Using an Intraoral Fluorescence Camera

Dental plaque accumulation is quantified using clinical indices or, otherwise, the planimetric plaque index (PPI),&#160;which measures the relative area of a tooth that is covered by plaque deposits. Compared to clinical indices, the PPI has a higher discriminatory power, but traditional planimetry is a time-consuming analysis, as the plaque-covered and clean tooth areas have to be determined manually for each image using image-processing software. Here, we present a method for the semi-automated planimetric quantification of dental plaque, which allows for the rapid processing of up to 1,000 images simultaneously. The method exploits the enhanced contrast between disclosed plaque, sound tooth surfaces, and soft tissues in fluorescence images acquired with an intraoral camera. Careful execution of the clinical procedures and accurate image acquisition are crucial steps for the successful semi-automated identification of the plaque-covered areas. The method is suitable for planimetry on sound facial and oral tooth surfaces, on most composite resin restorations, and on teeth with orthodontic brackets, but not on metallic restorations. Compared to traditional PPI recordings, semi-automated planimetry considerably reduces the amount of time spent on the analysis, as well as the subjective human input, thus increasing the reproducibility of planimetric measurements.

This study presents a semi-automated digital image analysis procedure for the planimetric quantification of disclosed dental plaque based on images acquired with an intraoral fluorescence camera. The method allows for the rapid and reliable quantification of dental plaque in the research environment.

使用口内荧光相机对牙菌斑进行半自动平面定量

Dental plaque accumulation is quantified using clinical indices or, otherwise, the planimetric plaque index (PPI),&#160;which measures the relative area ...

在白内障手术中通过 IOL 旋转预防后囊膜混浊

Rotating the Intraocular Lens to Prevent Posterior Capsular Opacification in Cataract Surgeries

Posterior capsule opacification (PCO) is a common postoperative complication of extracapsular cataract surgery, which is caused by the proliferation and migration of lens epithelial cells and can affect long-term visual outcomes significantly. The most effective treatment for PCO is neodymium-doped yttrium aluminum garnet (Nd:YAG) laser capsulotomy; however, this treatment is associated with posterior segment complication and can break the stability of capsular bag, affecting the position and function of trifocal or toric intraocular lenses (IOLs). Advances in surgical procedures, IOL design, and pharmacy have reduced the rate of PCO in recent years, concentrating on the inhibition of proliferative lens epithelial cells (LECs). This protocol aimed to clear LECs more thoroughly during phacoemulsification and IOL implantation. The first several steps, including clear corneal incision, continuous circular capsulorhexis, hydrodissection, hydrodelineation, and phacoemulsification, were completed as conventional procedures. After placing the IOL into the capsular bag, rotation of the IOL by at least 360&#176; was performed using an irrigation/aspiration tip or a hook, with slight stress on the posterior capsule. Some residuals occurred in the originally transparent capsular bag after rotation of the IOLs. Then, these materials and the viscoelastic were cleared completely using an irrigation/aspiration system. A clear posterior capsule was observed after the surgery in patients undergoing this method. This method of rotating IOLs is a simple, effective, and safe way to prevent PCO by clearing residual LECs and can be carried out without extra tools or skills.

作者聚焦：增强白内障手术的视觉效果：一种通过 IOL 旋转防止后囊膜混浊的新技术 

The present protocol describes removing residual epithelial cells by rotating the intraocular lens in extracapsular cataract surgery without extra tools for preventing posterior capsular opacification.

在白内障手术中旋转人工晶状体以防止后囊混浊

Posterior capsule opacification (PCO) is a common postoperative complication of extracapsular cataract surgery, which is caused by the proliferation and ...

用于临床规模 CAR-T 细胞生产的自动化处理器

Chimeric Antigen Receptor T Cell Manufacturing on an Automated Cell Processor

Chimeric antigen receptor (CAR)-T cells represent a promising immunotherapeutic approach for the treatment of various malignant and non-malignant diseases. CAR-T cells are genetically modified T cells that express a chimeric protein that recognizes and binds to a cell surface target, resulting in the killing of the target cell. Traditional CAR-T cell manufacturing methods are labor-intensive, expensive, and may carry the risk of contamination. The CliniMACS Prodigy, an automated cell processor, allows for manufacturing cell therapy products at a clinical scale in a closed system, minimizing the risk of contamination. Processing occurs semi-automatically under the control of a computer and thus&#160;minimizes human involvement in the process, which saves time and reduces variability and errors.
This manuscript and video describes the T cell transduction (TCT) process for manufacturing CAR-T cells using this processor. The TCT process involves CD4+/CD8+ T cell enrichment, activation, transduction with a viral vector, expansion, and harvest. Using the Activity Matrix, a functionality that allows ordering and timing of these steps, the TCT process can be customized extensively. We provide a walk-through of CAR-T cell manufacturing in compliance with current Good Manufacturing Practice&#160;(cGMP) and discuss required release testing and preclinical experiments that will support an Investigational New Drug (IND) application. We demonstrate&#160;the feasibility and discuss the advantages and disadvantages of using a&#160;semi-automatic process for clinical CAR-T cell manufacturing. Finally, we describe an ongoing investigator-initiated clinical trial that targets pediatric B-cell malignancies [NCT05480449] as an example of how this manufacturing process can be applied in a clinical setting.

作者聚焦：CAR-T 细胞制造和基因治疗生产的进步 

This article details the manufacturing process for chimeric antigen receptor T cells for clinical use, specifically using an automated cell processor capable of performing viral transduction and cultivation of T cells. We provide recommendations and describe pitfalls that should be considered during the process development and implementation of an early-phase clinical trial.