Name: medical-grade sterilizable target for fluid-immersed fetoscope optical distortion calibration
Uploaded: 2017-02-23T13:00:00.000+00:00
Duration: 7 min 3 s
Description: Watch this Scientific Journal Video about Medical-grade Sterilizable Target for Fluid-immersed Fetoscope Optical Distortion Calibration at JoVE.com

This work was supported through an Innovative Engineering for Health award by the Wellcome Trust [WT101957], the Engineering and Physical Sciences Research Council (EPSRC) [NS/A000027/1], and a National Institute for Health Research Biomedical Research Centre UCLH/UCL High Impact Initiative. Jan Deprest is being funded by the Fonds voor Wetenschappelijk Onderzoek Vlaanderen (FWO; JD as clinical researcher 1.8.012.07). Danail Stoyanov receives funding from the EPSRC (EP/N013220/1, EP/N022750/1), the EU-FP7 project CASCADE (FP7-ICT-2913-601021), and the EU-Horizon2020 project EndoVESPA (H2020-ICT- 2015-688592). Sebastien Ourselin receives funding from the EPSRC (EP/H046410/1, EP/J020990/1, EP/K005278) and the MRC (MR/J01107X/1). Marcel Tella is supported by the EPSRC-funded UCL Centre for Doctoral Training in Medical Imaging (EP/L016478/1).

1. Target Fabrication
<ol>
	<li>Sandblasting
	<ol>
		<li>Prepare a 316 stainless steel sheet with a 1.2-mm thickness. Using a pencil or a nail, draw a 40 mm x 40 mm square onto the sheet with the aid of a ruler.</li>
		<li>Cut the drawn square using a manual metal cutter. CAUTION! Watch the fingers.</li>
		<li>Use a file to round the corners and sides of the sample. CAUTION! They are very sharp; be careful.</li>
		<li>Prepare a straight wooden or metal block slightly larger in size than the stainless steel sheet. Place the cut sheet on it; do this in order to avoid bending the sample during sandblasting.</li>
		<li>Place the assembly in the internal blast chamber. Remember to use a dust collector and to tightly seal the internal blast chamber; otherwise, the sand will spread all over during the process. Wear safety goggles to protect the eyes.</li>
		<li>Position a blast gun perpendicular to and at least 4-5 cm away from the metal surface. Apply the foot control for sandblasting. Put the sample on the piece of wood (1-2 cm thick) using a vice, as the high-pressure sand flow can deform the sample. During sand-blasting, hold the sample tightly by the edge of the piece of wood or by using another vice.</li>
		<li>Repeat the sandblasting on the other side if it is desirable to have a calibration pattern engraved on both sides.</li>
	</ol>
	</li>
	<li>Laser patterning
	<ol>
		<li>Design a pattern of asymmetric circles, as shown in Figure 1.</li>
		<li>Prepare a drawing exchange format (DXF) file of the design either using CAD software or another suitable programming language. 
		NOTE: For convenience, a Python application that can generate DXF files for the design mentioned in this paper is provided as part of the compact GUI application16.</li>
		<li>Import the DXF files into the laser cutting software.</li>
		<li>Set up the following parameters for background etching. Laser Power: 40%, Scan Speed: 80 cm/s, Frequency: 4,000 Hz, Number of Passes: 1.</li>
		<li>Set up the following parameters for etching the pattern. Laser Power: 40%, Scan Speed: 2.1 cm/s, Frequency: 4,000 Hz, Number of Passes: 1.</li>
		<li>Put the sample on the working platform and align the cutting pattern using the software.</li>
		<li>After the laser performs the cut, clean the sample by dipping it in alcohol. Do not use any wipes, as they usually leave undesirable residue.</li>
	</ol>
	</li>
	<li>Sterilization
	<ol>
		<li>Wrap the sterilized sample in a sterilization package and insert it in the sterilization unit (autoclave).</li>
		<li>Add water (not distilled water) to the autoclave and follow the user&#39;s guide/manufacturer&#39;s recommendations to sterilize the target.</li>
	</ol>
	</li>
</ol>
2. Peri-operative Calibration
<ol>
	<li>Calibration software
	<ol>
		<li>Install the &#34;endocal&#34; endoscope calibration software package provided on GitHub16 (follow the instructions in the README file therein). 
		NOTE: This software wraps the OpenCV camera calibration module17 in an easy-to-use convenience application. The provided application runs in two modes: online and offline. The online mode acquires the video stream directly from compatible frame-grabber hardware. The offline mode allows for loading endoscope images either from a video file or a folder with a number of video frames saved as image files. See README for supported hardware and detailed instructions on how to use these two modes.</li>
	</ol>
	</li>
	<li>Endoscopic video acquisition 
	NOTE: The following instructions are for online calibration (as described above), but they are also applicable to offline calibration.
	<ol>
		<li>Place the calibration target in a sterile fluid container, such as a gallipot.</li>
		<li>Fill the container with the target fluid or a similar sterile substance. 
		NOTE: For instance, in fetoscopic procedures, the target fluid is amniotic fluid. Since the optical properties of amniotic fluid are similar to saline water18,19, sterile saline water can be used for calibrating the fetoscope.</li>
		<li>Adjust the zoom and sharpness of the endoscope as desired.</li>
		<li>Immerse the endoscope in the fluid and hold it at a distance from the calibration target similar to the distance from the anatomy that the endoscope will later be used at.</li>
		<li>Launch the calibration application and start the camera acquisition.</li>
		<li>Move the tip of the endoscope slightly for different views while keeping the whole calibration pattern in view of the camera. For optimal performance, keep the elliptical legend around the calibration pattern within the circular view of the endoscope. 
		NOTE: Video frames that are usable for calibration are indicated by a virtual pattern overlay, as seen in Figure 3.</li>
		<li>Acquire at least the minimum number of endoscopic camera views required for calibration (as indicated in the endocal window). 
		NOTE: The current version of endocal requires at least 10 endoscopic camera views for calibration, a heuristically selected number of views where the calibration error appears to be minimal and follow a stable pattern20.</li>
		<li>Press the calibration key, as indicated on the endocal window, to start the calibration process using the images acquired so far.</li>
	</ol>
	</li>
	<li>Saving and using the calibration parameters
	<ol>
		<li>Press the indicated calibration key to save the resulting calibration parameters in a YAML (&#34;YAML Ain&#39;t Markup Language&#34;) file21.</li>
		<li>Group the calibration parameters into the camera matrix and distortion coefficients, as explained in the OpenCV camera calibration module17. 
		NOTE: After performing the calibration, the calibration application automatically displays the distortion-corrected image to the right of the original endoscope image.</li>
		<li>Use the distortion-corrected video feed during a fetoscopic procedure for pure visualization or for real-time placental mosaicing11.</li>
	</ol>
	</li>
</ol>

<ol>
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Eng. 37 (8), 855-866 (1971).</li><li>Mallon, J., Whelan, P. 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A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fetal+surgery+is+a+clinical+reality.">Fetal surgery is a clinical reality.</a> Seminars in fetal and neonatal medicine. 15 (1), 58-67 (2009).</li><li>Watanabe, M., Flake, A. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fetal+surgery:+Progress+and+perspectives.">Fetal surgery: Progress and perspectives.</a> Advances in pediatrics. 57 (1), 353-372 (2010).</li><li>Lewi, L., Deprest, J., Hecher, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+vascular+anastomoses+in+monochorionic+twin+pregnancies+and+their+clinical+consequences.">The vascular anastomoses in monochorionic twin pregnancies and their clinical consequences.</a> American journal of obstetrics and gynecology. 208 (1), 19-30 (2013).</li><li>Yamashita, H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Miniature+bending+manipulator+for+fetoscopic+intrauterine+laser+therapy+to+treat+twin-to-twin+transfusion+syndrome.">Miniature bending manipulator for fetoscopic intrauterine laser therapy to treat twin-to-twin transfusion syndrome.</a> Surgical Endoscopy. 22 (2), 430-435 (2008).</li><li>Daga, P., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Real-time+mosaicing+of+fetoscopic+videos+using+SIFT.">Real-time mosaicing of fetoscopic videos using SIFT.</a> Proc. SPIE 9786, Medical Imaging 2016: Image-Guided Procedures, Robotic Interventions, and Modeling. 97861R. , (2016).</li><li>Yang, L., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Image+mapping+of+untracked+free-hand+endoscopic+views+to+an+ultrasound+image-constructed+3D+placenta+model.">Image mapping of untracked free-hand endoscopic views to an ultrasound image-constructed 3D placenta model.</a> The International Journal of Medical Robotics and Computer Assisted Surgery. 11 (2), 223-234 (2015).</li><li>Liao, H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fast+image+mapping+of+endoscopic+image+mosaics+with+three-dimensional+ultrasound+image+for+intrauterine+fetal+surgery.">Fast image mapping of endoscopic image mosaics with three-dimensional ultrasound image for intrauterine fetal surgery.</a> Minimally invasive therapy &amp; allied technologies. 18 (6), 332-340 (2009).</li><li>Chadebecq, F., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Practical+Dry+Calibration+With+Medium+Adaptation+For+Fluid-Immersed+Endoscopy.">Practical Dry Calibration With Medium Adaptation For Fluid-Immersed Endoscopy.</a> , (2015).</li><li>Wengert, C., Reeff, M., Cattin, P. C., Sz&#233;kely, G., Handels, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Bildverarbeitung f&#252;r die Medizin 2006: Algorithmen Systeme Anwendungen. Proceedings des Workshops vom 19. - 21. M&#228;rz 2006 in Hamburg. , 419-423 (2006).</li><li>. Compact GUI application for optical distortion calibration of endoscopes Available from: <a target="_blank" href="https://github.com/gift-surg/endocal">https://github.com/gift-surg/endocal</a> (2016)</li><li>Reeff, M., Gerhard, F., Cattin, P. C., Sz&#233;kely, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Mosaicing of endoscopic placenta images. , (2011).</li><li>Steigman, S. A., Kunisaki, S. M., Wilkins-Haug, L., Takoudes, T. C., Fauza, D. O. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Optical+properties+of+human+amniotic+fluid:+implications+for+videofetoscopic+surgery.">Optical properties of human amniotic fluid: implications for videofetoscopic surgery.</a> Fetal diagnosis and therapy. 27 (2), 87-90 (2009).</li><li>Zhang, Z. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+flexible+new+technique+for+camera+calibration.">A flexible new technique for camera calibration.</a> IEEE Transactions on Pattern Analysis and Machine Intelligence. 22 (11), 1330-1334 (2000).</li><li>. Open Source Computer Vision Library Available from: <a target="_blank" href="https://github.com/opencv/opencv">https://github.com/opencv/opencv</a> (2016)</li></ol>

The authors have nothing to disclose.

Sandblasting is an important step in the fabrication process because the raw metal surface prominently reflects the endoscope light, making it impossible for the circles to be detected. It is difficult to distinguish the circles even with the naked eye (see Figure 5). Note that the surface of the target shown was already etched with a laser. However, this does not diminish light reflection.
<img alt="Figure 5" src="/files/ftp_upload/55298/55298fig5.jpg" /> 
Figure 5: Calibration target with no sandblasting applied. As seen from the endoscope view on the left, the glare from the endoscope light on the material surface makes it difficult even for the naked eye to distinguish the circles (there is a circle just to the southeast of the large reflection). Note that the surface of this target (i.e., the &#34;background&#34;) was already etched, but this is not helpful in the absence of sandblasting. <a href="http://cloudflare.jove.com/files/ftp_upload/55298/55298fig5large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
Prior to pattern etching, it is also important to etch the surface of the whole sample. This is necessary because the sandblasted surface has many specular reflections (see Figure 6), which interfere with blob detection.
<img alt="Figure 6" src="/files/ftp_upload/55298/55298fig6.jpg" /> 
Figure 6: Sandblasted surface with no etching. Although not as prominent as the raw metal surface, the relatively small specular reflections (some of which are highlighted with yellow arrows) are still sufficient to prevent blob detection from succeeding, so no calibration can be performed with this target. <a href="http://cloudflare.jove.com/files/ftp_upload/55298/55298fig6large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
Applying the laser at different speeds gives different background colors. The background color plays a significant role in the contrast between the circles and the background. Hence, it is vital to determine the optimal background color. For this purpose, a plate with circles etched against a set of different backgrounds was created (see Figure 7). The backgrounds were tested using the feature detection module of OpenCV23, which is used in the OpenCV camera calibration module17. In this work, the target was made of stainless steel, as it is the most common and reliable material used in clinics for medical devices. This material is freely available, not expensive, robust, and easy to sterilize. Other materials could potentially be used for the calibration target, such as aluminum or iodized metals, but this is the scope of future work.
<img alt="Figure 7" src="/files/ftp_upload/55298/55298fig7.jpg" /> 
Figure 7: Stainless steel plate featuring a palette of different background colors etched with the laser. Practical experiments were conducted in conjunction with the OpenCV feature detection module to determine which background color gives the optimal result in terms of blob-to-background contrast23. The endoscope view on the left shows the plate. The moderate background colors (i.e., those other that the darkest and lightest ones) in this palette yield better blob detection. <a href="http://cloudflare.jove.com/files/ftp_upload/55298/55298fig7large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
One of the advantages of this work is that performing a calibration using the fabricated target takes 2-3 min. Most of the effort goes to manually stabilizing the endoscope to obtain decent views of the calibration pattern. Using a custom-built endoscope holder could eliminate the need for manual stabilization, which in turn could significantly shorten calibration time.
<img alt="Video 1" src="/files/ftp_upload/55298/55298video1.jpg" /> 
Video 1: Video showing how optical distortion calibration can be performed using the developed calibration target together with the endocal software. <a href="http://cloudflare.jove.com/files/ftp_upload/55298/fetoscope-calibration-480.mov" target="_blank">Please click here to view this video. (Right-click to download.)</a>
An advantage of our work compared to the work of Wengert et al.15 is that the OpenCV camera calibration module17 can be used as is for calibration, without requiring any modification or custom parameterization. Because OpenCV is a well-established and well-maintained software package and is very popular in the computer vision community, using it eliminates the need for writing and maintaining custom software. For the reader&#39;s convenience, a compact GUI application is provided16, which the reader can easily install and use to test new calibration targets. One disadvantage of our method compared to Wengert et al.15 is that their method is more robust to occlusions of the pattern, as it does not require the detection of all blobs.
Initially, a calibration target with a checkerboard pattern was fabricated for this work. However, this type of calibration target proved to be unsuitable in experiments due to the difficulty of detecting the corners of the checkerboard squares. Corner detection relies on histogram-based image binarization (see the OpenCV source code24). This implies the need for a clear color contrast between the dark and light squares, which could not be guaranteed with our checkerboard pattern, partially due to specular reflections, like the ones shown in Figure 6. Such specular reflections are present even after background etching; however, the detection of the circles seems to be less sensitive to this shortcoming.
In the current setup, only perpendicular views of the calibration target allow for successful blob detection. This is due to the specular reflections from the target surface hampering blob detection at oblique angles. We are working to further improve the target so as to allow for the acquisition of views at a wider range of angles, which could potentially improve the quality of performed calibrations20.
In the real-time placental mosaicing pipeline that was previously proposed11, the computation of the transformation that maps image pairs relies on the successful detection and grouping of features. Optical distortions, on the other hand, cause a group of features with a rigid geometry to appear different across images. As a consequence, this difference leads to inaccuracies in the computed transformations, which cause drifts in the resulting image mosaics. Because the most prominent optical distortions are present towards the edges, endoscopic images are currently cropped to their innermost regions. A good correction for optical distortions would potentially allow for the incorporation of a larger part of each image into the mosaicing process. The advantage of this method is two-fold. First, it would increase the number of detected features in each image, potentially improving the computation of the image transformations. Second, it would allow for the whole target anatomical surface to be reconstructed in a shorter time.

Camera calibration is a well-known problem in the computer vision field that has been intensively studied over the years1,2,3. A key step of camera calibration procedures is to estimate the parameters of a distortion model, as well as the intrinsic camera parameters, by extracting a grid of points with a known geometry from camera images with sub-pixel accuracy. Calibration targets with a checkerboard pattern featuring black and white squares are commonly used for this purpose. Circular blobs offer an alternative pattern4,5,6.
In recent years, there has been a growing interest in the development of surgical navigation technology for fetal surgery procedures, such as the treatment of twin-to-twin transfusion syndrome (TTTS) on fetuses7,8,9,10. As the field of view of the fetoscope (i.e., an endoscope used in fetal surgical procedures) is very limited, methods for mapping the placental vasculature without the use of external trackers have been proposed to aid TTTS surgery11,12,13. Optical distortions within fetoscopic images have adverse effects on these computational mosaicing methods that rely on visual information extraction11. Thus, there is an unmet need for a cost-efficient and fast tool for peri-operatively calibrating fetoscopes so that optical distortion compensation can be done in real-time during the intervention.
Due to the fact that the fetoscope is immersed in amniotic fluid during the intervention, the refraction index difference between air and amniotic fluid renders classical in-air camera calibration methods unsuitable for fetal surgery procedures. Estimating fluid-immersed camera parameters from in-air camera parameters is a difficult task and requires at least one image of the fluid-immersed calibration target14. Furthermore, peri-operative, fluid-immersed fetoscopic camera calibration is currently impractical due to sterilization requirements and restrictions on the materials allowed in the operating theater. Due to these reasons, calibrating endoscopes for optical distortions is typically not part of the current clinical workflow. The work in this manuscript is an attempt to close this camera calibration gap by designing and producing a sterilizable and practical optical distortion calibration target featuring a pattern of asymmetric circles. Previously, Wengert et al. fabricated a custom calibration device featuring an oxidized aluminum plate as the calibration target. Their method, however, works only in conjunction with the custom calibration algorithm they developed15.

<table><tbody><tr><td>1.2 mm Metal sheet 316 Grade, 40 mm by 40 mm</td><td></td><td></td><td></td></tr><tr><td>Water container at least 50 mm by 50 mm by 30 mm</td><td></td><td></td><td></td></tr><tr><td>A sterilization package</td><td></td><td></td><td></td></tr><tr><td>Saline water</td><td></td><td></td><td></td></tr><tr><td>Manual metal cutter</td><td></td><td></td><td></td></tr><tr><td>A file to round up the corners</td><td></td><td></td><td></td></tr><tr><td>A wooden or metal block 50 mm by 50 mm at least 10 mm thick</td><td></td><td></td><td></td></tr><tr><td>A vice (desirable but not required)</td><td></td><td></td><td></td></tr><tr><td>Sand Blasting machine</td><td></td><td></td><td></td></tr><tr><td>GUI application to create .dxf file with the pattern</td><td></td><td></td><td>https://github.com/gift-surg/endocal</td></tr><tr><td>PC</td><td></td><td></td><td></td></tr><tr><td>Laser Cutter</td><td></td><td></td><td></td></tr><tr><td>Autoclave</td><td></td><td></td><td></td></tr><tr><td>An endoscope calibration software</td><td></td><td></td><td>from GitHub: https://github.com/gift-surg/endocal</td></tr><tr><td>Endoscope</td><td></td><td></td><td></td></tr><tr><td>OpenCV camera calibration module</td><td></td><td></td><td>http://docs.opencv.org/2.4/doc/tutorials/calib3d/camera_calibration/camera_calibration.html</td></tr><tr><td>Safety goggles</td><td></td><td></td><td></td></tr><tr><td>A lab coat</td><td></td><td></td><td></td></tr><tr><td>A ruler and a marker</td><td></td><td></td><td></td></tr><tr><td>Alcohol (preferably ethanol) for dust removal and cleaning</td><td></td><td></td><td></td></tr></tbody></table>

medical-grade sterilizable target for fluid-immersed fetoscope optical distortion calibration

We have developed a calibration target for use with fluid-immersed endoscopes within the context of the GIFT-Surg (Guided Instrumentation for Fetal Therapy and Surgery) project. One of the aims of this project is to engineer novel, real-time image processing methods for intra-operative use in the treatment of congenital birth defects, such as spina bifida and the twin-to-twin transfusion syndrome. The developed target allows for the sterility-preserving optical distortion calibration of endoscopes within a few minutes. Good optical distortion calibration and compensation are important for mitigating undesirable effects like radial distortions, which not only hamper accurate imaging using existing endoscopic technology during fetal surgery, but also make acquired images less suitable for potentially very useful image computing applications, like real-time mosaicing. In this paper proposes a novel fabrication method to create an affordable, sterilizable calibration target suitable for use in a clinical setup. This method involves etching a calibration pattern by laser cutting a sandblasted stainless steel sheet. This target was validated using the camera calibration module provided by OpenCV, a state-of-the-art software library popular in the computer vision community.

We created a sterilizable calibration target by etching a pattern of asymmetric circle on a sandblasted stainless steel metal sheet, whose design is shown in Figure 1. An exemplar setup showing this calibration target in action together with a fetoscope is shown in Figure 2. To feed this design into the laser etching software, a custom application was implemented in the Python programming language16. Creating the design pattern involves iteratively etching parallel lines on a metal sheet. For the pattern to have a consistent color in the end, the distance between these lines should be less than the width of the laser beam (see the inset of Figure 1)&#8212;this value is 45 &#181;m for the Violino (Laservall) laser cutter.
<img alt="Figure 1" src="/files/ftp_upload/55298/55298fig1.jpg" /> 
Figure 1: Design of the engraved pattern featuring a 3-by-11 grid of asymmetric circles. Inset: zoomed-in view of the grid of asymmetric circles. The distance between the lines is 45 &#181;m (equal to the laser beam width), and each circle has a diameter of 1 mm. Other sizes could be used for the grid as well, but this was found to be optimal with respect to the fetoscope field of view. <a href="http://cloudflare.jove.com/files/ftp_upload/55298/55298fig1large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 2" src="/files/ftp_upload/55298/55298fig2.jpg" /> 
Figure 2: Exemplar setup with the calibration target in use. The tip of the water-immersed fetoscope is directed at the calibration target on the right. On the left is a British penny to provide scale information. <a href="http://cloudflare.jove.com/files/ftp_upload/55298/55298fig2large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
The fabricated calibration target allows for the detection of the circular pattern in the endoscopic video stream with OpenCV17, whose locations are then sorted into the pre-defined asymmetric circular grid (see Figure 3). Using this information in conjunction with the already-known grid geometry, internal camera parameters can be estimated. These include the camera matrix and the distortion coefficients. The camera matrix consists of the focal lengths and the optical centers along the x- and y-axis of the 2D image plane. The distortion coefficients are based on the Brown-Conrady model3. Note that for this work, only the radial distortion parameters were estimated. For a brief discussion of the theory, with practical examples, see the webpage of the OpenCV camera calibration module17 and the MATLAB camera calibration toolbox22. More details about the camera calibration procedure are available in Zhang&#39;s work20. The endocal software repository features a sample dataset of 10 endoscopic views of the fabricated calibration target16. Using this dataset, a calibration with an average re-projection error of 0.28 pixels (min: 0.16, max: 0.45) was obtained. This is comparable to the 0.25 pixels reported by Wengert et al. using their custom calibration algorithm15. The same research group, however, reported a re-projection error of 0.6 pixels in a more recent paper when using the method in15 for calibrating an endoscopic camera used for placental mosaicing18.
<img alt="Figure 3" src="/files/ftp_upload/55298/55298fig3.jpg" /> 
Figure 3: Real-time detection of the calibration pattern. A screenshot from the calibration application16 featuring the detected calibration pattern overlaid onto the live video stream using the virtual reality visualization from OpenCV17. Note that each detected column of the calibration pattern is emphasized by a different color. The detected circles, in conjunction with the known geometry, are used for computing the camera parameters. <a href="http://cloudflare.jove.com/files/ftp_upload/55298/55298fig3large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
The estimated camera parameters are used for optical distortion correction. Figure 4 shows a rectangular chessboard pattern, as viewed using a fetoscope, where optical distortions make the lines appear as curves. Note that the lines appear normal in the distortion-corrected image.
<img alt="Figure 4" src="/files/ftp_upload/55298/55298fig4.jpg" /> 
Figure 4: Optical distortion correction. A screenshot from the calibration application16 featuring the live video image from a fetoscope recording from the checkerboard pattern (left) with the distortion-corrected image (right). Three exemplary lines are drawn in both images, each from one corner to another, where the trajectory is linear. Due to the optical distortions, these lines appear as curves in the original fetoscope images. <a href="http://cloudflare.jove.com/files/ftp_upload/55298/55298fig4large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>

Watch this Scientific Journal Video about Medical-grade Sterilizable Target for Fluid-immersed Fetoscope Optical Distortion Calibration at JoVE.com

Medical-grade Sterilizable Target for Fluid-immersed Fetoscope Optical Distortion Calibration

This article describes the design and development of a sterilizable custom camera optical distortion calibration target for the peri-operative, fluid-immersed calibration of endoscopes during endoscopic interventions.

We have developed a calibration target for use with fluid-immersed endoscopes within the context of the GIFT-Surg (Guided Instrumentation for Fetal ...

medical-grade-sterilizable-target-for-fluid-immersed-fetoscope

Research

JoVE Journal

Bioengineering

7.6K Views.  University College London. This article describes the design and development of a sterilizable custom camera optical distortion calibration target for the peri-operative, fluid-immersed calibration of endoscopes during endoscopic interventions.

Video: Medical-grade Sterilizable Target for Fluid-immersed Fetoscope Optical Distortion Calibration

Evaluation of the Feasibility, Safety, and Accuracy of an Intraoperative High-intensity Focused Ultrasound Device for Treating Liver Metastases

Today, the only potentially curative option in patients with colorectal liver metastases is surgery. However, liver resection is feasible in less than 20% of patients. Surgery has been widely used in association with radiofrequency, cryotherapy, or microwaves to expand the number of treatments performed with a curative intent. Nevertheless, several limitations have been documented when using these techniques (i.e., a traumatic puncture of the parenchyma, a limited size of lesions, and an inability to monitor the treatment in real-time).High-intensity focused ultrasound (HIFU) technology can achieve precise ablations of biological tissues without incisions or radiation. Current HIFU devices are based on an extracorporeal approach with limited access to the liver. We have developed a HIFU device designed for intraoperative use. The use of a toroidal transducer enables an ablation rate (10 cm3&#183;min-1) higher than any other treatment and is independent of perfusion.
The feasibility, safety, and accuracy of intraoperative HIFU ablation were evaluated during an ablate-and-resect prospective study. This clinical phase I and IIa study was performed in patients undergoing hepatectomy for liver metastases. The HIFU treatment was performed on healthy tissue scheduled for resection.Liver metastases measuring less than 20 mm will be targeted during phase IIb (currently ongoing). This set-up allows the real-time evaluation of HIFU ablation while protecting participating patients from any adverse effects related to this new technique.
Fifteen patients were included in phase I&#8211;IIa and 30 HIFU ablations were safely created within 40 s and with a precision of 1&#8211;2 mm. The average dimensions of the HIFU ablations were 27.5 x 21.0 mm2, corresponding to a volume of approximately 7.5 cm3. The aim of the ongoing phase IIb is to ablate metastases of less than 20 mm in diameter with a 5 mm margin.

Here, we present an ablate-and-resect prospective study to evaluate the feasibility, safety, and accuracy of intraoperative high-intensity focused ultrasound ablation in patients undergoing hepatectomy for liver metastases.

Today, the only potentially curative option in patients with colorectal liver metastases is surgery. However, liver resection is feasible in less than 20% ...

Cancer Research

Three-Dimensional Ultrasonic Needle Tip Tracking with a Fiber-Optic Ultrasound Receiver

Ultrasound is frequently used for guiding minimally invasive procedures, but visualizing medical devices is often challenging with this imaging modality. When visualization is lost, the medical device can cause trauma to critical tissue structures. Here, a method to track the needle tip during ultrasound image-guided procedures is presented. This method involves the use of a fiber-optic ultrasound receiver that is affixed within the cannula of a medical needle to communicate ultrasonically with the external ultrasound probe. This custom probe comprises a central transducer element array and side element arrays. In addition to conventional two-dimensional (2D) B-mode ultrasound imaging provided by the central array, three-dimensional (3D) needle tip tracking is provided by the side arrays. For B-mode ultrasound imaging, a standard transmit-receive sequence with electronic beamforming is performed. For ultrasonic tracking, Golay-coded ultrasound transmissions from the 4 side arrays are received by the hydrophone sensor, and subsequently the received signals are decoded to identify the needle tip&#39;s spatial location with respect to the ultrasound imaging probe. As a preliminary validation of this method, insertions of the needle/hydrophone pair were performed in clinically realistic contexts. This novel ultrasound imaging/tracking method is compatible with current clinical workflow, and it provides reliable device tracking during in-plane and out-of-plane needle insertions.

Accurate and efficient visualization of invasive medical devices is extremely important in many ultrasound-guided minimally invasive procedures. Here, a method for localizing the spatial position of a needle tip relative to the ultrasound imaging probe is presented.

Ultrasound is frequently used for guiding minimally invasive procedures, but visualizing medical devices is often challenging with this imaging modality. ...

Engineering

Fabrication of Refractive-index-matched Devices for Biomedical Microfluidics

The use of microfluidic devices has emerged as a defining tool for biomedical applications. When combined with modern microscopy techniques, these devices can be implemented as part of a robust platform capable of making simultaneous complementary measurements. The primary challenge created by the combination of these two techniques is the mismatch in refractive index between the materials traditionally used to make microfluidic devices and the aqueous solutions typically used in biomedicine. This mismatch can create optical artifacts near the channel or device edges. One solution is to reduce the refractive index of the material used to fabricate the device by using a fluorinated polymer such as MY133-V2000 whose refractive index is similar to that of water (n = 1.33). Here, the construction of a microfluidic device made out of MY133-V2000 using soft lithography techniques is demonstrated, using O2 plasma in conjunction with an acrylic holder to increase the adhesion between the MY133-V2000 fabricated device and the polydimethylsiloxane (PDMS) substrate. The device is then tested by incubating it filled with cell culture media for 24 h to demonstrate the ability of the device to maintain cell culture conditions during the course of a typical imaging experiment. Finally, quantitative phase microscopy (QPM) is used to measure the distribution of mass within the live adherent cells in the microchannel. This way, the increased precision, enabled by fabricating the device from a low index of refraction polymer such as MY133-V2000 in lieu of traditional soft lithography materials such as PDMS, is demonstrated. Overall, this approach for fabricating microfluidic devices can be readily integrated into existing soft lithography workflows in order to reduce optical artifacts and increase measurement precision.

This protocol describes the fabrication of microfluidic devices from MY133-V2000 to eliminate artifacts that often arise in microchannels due to the mismatching refractive indices between microchannel structures and an aqueous solution. This protocol uses an acrylic holder to compress the encapsulated device, improving adhesion both chemically and mechanically.

The use of microfluidic devices has emerged as a defining tool for biomedical applications. When combined with modern microscopy techniques, these devices ...

Multiplexing Focused Ultrasound Stimulation with Fluorescence Microscopy

By focusing low-intensity ultrasound pulses that penetrate soft tissues, LIPUS represents a promising biomedical technology to remotely and safely manipulate neural firing, hormonal secretion and genetically-reprogrammed cells. However, the translation of this technology for medical applications is currently hampered by a lack of biophysical mechanisms by which targeted tissues sense and respond to LIPUS. A suitable approach to identify these mechanisms would be to use optical biosensors in combination with LIPUS to determine underlying signaling pathways. However, implementing LIPUS to a fluorescence microscope may introduce undesired mechanical artefacts due to the presence of physical interfaces that reflect, absorb and refract acoustic waves. This article presents a step-by-step procedure to incorporate LIPUS to commercially-available upright epi-fluorescence microscopes while minimizing the influence of physical interfaces along the acoustic path. A simple procedure is described to operate a single-element ultrasound transducer and to bring the focal zone of the transducer into the objective focal point. The use of LIPUS is illustrated with an example of LIPUS-induced calcium transients in cultured human glioblastoma cells measured using calcium imaging.

Low-Intensity Pulsed Ultrasound Stimulation (LIPUS) is a modality for non-invasive mechanical stimulation of endogenous or engineered cells with high spatial and temporal resolution. This article describes how to implement LIPUS to an epi-fluorescence microscope and how to minimize acoustic impedance mismatch along the ultrasound path to prevent unwanted mechanical artefacts.

By focusing low-intensity ultrasound pulses that penetrate soft tissues, LIPUS represents a promising biomedical technology to remotely and safely ...

Evaluating Targeting Accuracy in the Focal Plane for an Ultrasound-guided High-intensity Focused Ultrasound Phased-array System

Phased arrays are increasingly used as high-intensity focused ultrasound (HIFU) transducers in the existing extracorporeal ultrasound-guided HIFU (USgHIFU) systems. The HIFU transducers in such systems are usually spherical in shape with a central hole where a US imaging probe is mounted and can be rotated. The image on the plane of treatment can be reconstructed through the image sequence acquired during the rotation of the probe. Therefore, the treatment plan can be made on the reconstructed images. In order to evaluate the targeting accuracy in the focal plane of such systems, the protocol of a method using a bovine muscle and marker-embedded phantom is described. In the phantom, four solid balls at the corners of a square resin model serve as the reference markers in the reconstructed image. The target should be moved so that both its center and the center of the square model can coincide according to their relative positions in the reconstructed image. Swine muscle with a thickness of about 30 mm is placed above the phantom to mimic the beam path in clinical settings. After sonication, the treatment plane in the phantom is scanned and the boundary of the associated lesion is extracted from the scanned image. The targeting accuracy can be evaluated by measuring the distance between the centers of target and lesion, as well as three derivative parameters. This method cannot only evaluate the targeting accuracy of the target consisting of multiple focal spots rather than a single focal spot in a clinically relevant beam path of the USgHIFU phased-array system, but it can be also used in the preclinical evaluation or regular maintenance of USgHIFU systems configured with phased-array or self-focused HIFU transducer.

This study describes a protocol to evaluate the targeting accuracy in the focal plane of an ultrasound-guided high-intensity focused ultrasound phased-array system.

Phased arrays are increasingly used as high-intensity focused ultrasound (HIFU) transducers in the existing extracorporeal ultrasound-guided HIFU ...

Microtensiometer for Confocal Microscopy Visualization of Dynamic Interfaces

Adsorption of surface-active molecules to fluid-fluid interfaces is ubiquitous in nature. Characterizing these interfaces requires measuring surfactant adsorption rates, evaluating equilibrium surface tensions as a function of bulk surfactant concentration, and relating how surface tension changes with changes in the interfacial area following equilibration. Simultaneous visualization of the interface using fluorescence imaging with a high-speed confocal microscope allows the direct evaluation of structure-function relationships. In the capillary pressure microtensiometer (CPM), a hemispherical air bubble is pinned at the end of the capillary in a 1 mL volume liquid reservoir. The capillary pressure across the bubble interface is controlled via a commercial microfluidic flow controller that allows for model-based pressure, bubble curvature, or bubble area control based on the Laplace equation. Compared to previous techniques such as the Langmuir trough and pendant drop, the measurement and control precision and response time are greatly enhanced; capillary pressure variations can be applied and controlled in milliseconds. The dynamic response of the bubble interface is visualized via a second optical lens as the bubble expands and contracts. The bubble contour is fit to a circular profile to determine the bubble curvature radius, R, as well as any deviations from circularity that would invalidate the results. The Laplace equation is used to determine the dynamic surface tension of the interface. Following equilibration, small pressure oscillations can be imposed by the computer-controlled microfluidic pump to oscillate the bubble radius (frequencies of 0.001-100 cycles/min) to determine the dilatational modulus The overall dimensions of the system are sufficiently small that the microtensiometer fits under the lens of a high-speed confocal microscope allowing fluorescently tagged chemical species to be quantitatively tracked with submicron lateral resolution.

This manuscript describes the design and operation of a microtensiometer/confocal microscope to do simultaneous measurements of interfacial tension and surface dilatational rheology while visualizing the interfacial morphology. This provides the real-time construction of structure-property relationships of interfaces important in technology and physiology.

Adsorption of surface-active molecules to fluid-fluid interfaces is ubiquitous in nature. Characterizing these interfaces requires measuring surfactant ...

Direct Force Measurements of Subcellular Mechanics in Confinement using Optical Tweezers

During the development of a multicellular organism, a single fertilized cell divides and gives rise to multiple tissues with diverse functions. Tissue morphogenesis goes in hand with molecular and structural changes at the single cell level that result in variations of subcellular mechanical properties. As a consequence, even within the same cell, different organelles and compartments resist differently to mechanical stresses; and mechanotransduction pathways can actively regulate their mechanical properties. The ability of a cell to adapt to the microenvironment of the tissue niche thus is in part due to the ability to sense and respond to mechanical stresses. We recently proposed a new mechanosensation paradigm in which nuclear deformation and positioning enables a cell to gauge the physical 3D environment and endows the cell with a sense of proprioception to decode changes in cell shape. In this article, we describe a new method to measure the forces and material properties that shape the cell nucleus inside living cells, exemplified on adherent cells and mechanically confined cells. The measurements can be performed non-invasively with optical traps inside cells, and the forces are directly accessible through calibration-free detection of light momentum. This allows measuring the mechanics of the nucleus independently from cell surface deformations and allowing dissection of exteroceptive and interoceptive mechanotransduction pathways. Importantly, the trapping experiment can be combined with optical microscopy to investigate the cellular response and subcellular dynamics using fluorescence imaging of the cytoskeleton, calcium ions, or nuclear morphology. The presented method is straightforward to apply, compatible with commercial solutions for force measurements, and can easily be extended to investigate the mechanics of other subcellular compartments, e.g., mitochondria, stress-fibers, and endosomes.

Here, we present a protocol to investigate the intracellular mechanical properties of isolated embryonic zebrafish cells in three-dimensional confinement with direct force measurement by an optical trap.

During the development of a multicellular organism, a single fertilized cell divides and gives rise to multiple tissues with diverse functions. Tissue ...

Developmental Biology

Spatio-Temporal In Vivo Imaging of Ocular Drug Delivery Systems using Fiberoptic Confocal Laser Microendoscopy

Subconjunctival injection is an attractive route to administer ocular drugs due to easy trans-scleral access that bypasses anterior ocular barriers, such as the cornea and conjunctiva. While therapeutic effects and pharmacokinetics of the drugs upon subconjunctival injection have been described in some studies, very few assess the ocular distribution of drugs or drug delivery systems (DDS). The latter is critical for the optimization of intraocular DDS design and drug bioavailability to achieve the desired ocular localization and duration of action (e.g., acute versus. prolonged). This study establishes the use of fiberoptic confocal laser microendoscopy (CLM) to qualitatively study the ocular distribution of fluorescent liposomes in real-time in live mice after sub-conjunctival injection. Being designed for in vivo visual inspection of tissues at the microscopic level, this is also the first full description of the CLM imaging method to study spatio-temporal distribution of injectables in the eye after subconjunctival injection.

Spatio-Temporal In Vivo Imaging of Ocular Drug Delivery Systems using Fiberoptic Confocal Laser Microendoscopy

We present a protocol for the use of fiberoptic confocal laser microendoscopy (CLM) to non-invasively study the spatio-temporal distribution of liposomes in the eye after subconjunctival injection.

Subconjunctival injection is an attractive route to administer ocular drugs due to easy trans-scleral access that bypasses anterior ocular barriers, such ...

Model Surgical Training: Skills Acquisition in Fetoscopic Laser Photocoagulation of Monochorionic Diamniotic Twin Placenta Using Realistic Simulators

Fetoscopic laser coagulation of arterio-venous anastomoses (AVA) in a monochorionic placenta is the standard of care for twin-twin transfusion syndrome (TTTS), but is technically challenging and can lead to significant complications. Acquiring and maintaining the necessary surgical skills require consistent practice, a critical caseload, and time. Training on realistic surgical simulators can potentially shorten this steep learning curve and enables several proceduralists to acquire procedure-specific skills simultaneously. Here we describe realistic simulators designed to allow the user familiarity with the equipment and specific steps required in the surgical treatment of TTTS, including fetoscopic handling, approaches to anterior and posterior placenta, recognition of anastomoses, and efficient coagulation of vessels. We describe the skills that are especially important in conducting placental laser coagulation that the surgeon can practice on the model and apply in a clinical case. These models can be adapted easily depending on the availability of materials and require standard fetoscopy equipment. Such training systems are complementary to traditional surgical apprenticeships and can be useful aids for fetal medicine units that provide this clinical service.

Practicing the specific skills required for fetoscopic laser coagulation of monochorionic placental anastomoses on realistic models can aid less experienced surgeons in overcoming the steep learning curve associated with this procedure that is now regarded as the standard of care for twin-twin transfusion syndrome.

Fetoscopic laser coagulation of arterio-venous anastomoses (AVA) in a monochorionic placenta is the standard of care for twin-twin transfusion syndrome ...

Medicine

Biomechanical Characterization of Porcine Ocular Lens with Automated Compression

Automated Compression Testing of the Ocular Lens

The biomechanical properties of the ocular lens are essential to its function as a variable power optical element. These properties change dramatically with age in the human lens, resulting in a loss of near vision called presbyopia. However, the mechanisms of these changes remain unknown. Lens compression offers a relatively simple method for assessing the lens' biomechanical stiffness in a qualitative sense and, when coupled with appropriate analytical techniques, can help quantify biomechanical properties. A variety of lens compression tests have been performed to date, including both manual and automated, but these methods inconsistently apply key aspects of biomechanical testing such as preconditioning, loading rates, and time between measurements. This paper describes a fully automated lens compression test wherein a motorized stage is synchronized with a camera to capture the force, displacement, and shape of the lens throughout a preprogrammed loading protocol. A characteristic elastic modulus may then be calculated from these data. While demonstrated here using porcine lenses, the approach is appropriate for the compression of lenses of any species.

Author Spotlight: Advancing Understanding of Age-Related Lens Stiffness Changes 

We present an automated method for characterizing the effective elastic modulus of an ocular lens using a compression test.

The biomechanical properties of the ocular lens are essential to its function as a variable power optical element. These properties change dramatically ...

Medical-grade Sterilizable Target for Fluid-immersed Fetoscope Optical Distortion Calibration

Summary

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Chapters in this video

Evaluation of the Feasibility, Safety, and Accuracy of an Intraoperative High-intensity Focused Ultrasound Device for Treating Liver Metastases

Three-Dimensional Ultrasonic Needle Tip Tracking with a Fiber-Optic Ultrasound Receiver

Fabrication of Refractive-index-matched Devices for Biomedical Microfluidics

Multiplexing Focused Ultrasound Stimulation with Fluorescence Microscopy

Evaluating Targeting Accuracy in the Focal Plane for an Ultrasound-guided High-intensity Focused Ultrasound Phased-array System

Microtensiometer for Confocal Microscopy Visualization of Dynamic Interfaces

Direct Force Measurements of Subcellular Mechanics in Confinement using Optical Tweezers

Spatio-Temporal In Vivo Imaging of Ocular Drug Delivery Systems using Fiberoptic Confocal Laser Microendoscopy

Model Surgical Training: Skills Acquisition in Fetoscopic Laser Photocoagulation of Monochorionic Diamniotic Twin Placenta Using Realistic Simulators

Automated Compression Testing of the Ocular Lens