This project was funded by the Simulation Training Center (STC) at the University of California, San Diego School of Medicine in La Jolla, CA. We would like to thank Blake Freechtle for his contributions to Figure 5.

For this project, authors JR and PS volunteered as ultrasound subjects, and verbal consent was obtained from both. For those following this protocol, obtain approval from an ethics committee or institutional review board (IRB) prior to using patients or human volunteers as study subjects.
1. Phantom design and silicone mold creation
<ol>
	<li>Creation of a reference ultrasound image
	<ol>
		<li>For each ultrasound phantom, have a physician with ultrasound sub-specialization and familiarity with the nerve block procedure being simulated by the desired phantom create a reference ultrasound image from a volunteer human subject (Figure 1). Ensure that this ultrasound image has a view transverse to the applicable nerve or tissue plane into which the anesthetic would be injected.</li>
	</ol>
	</li>
	<li>Design and 3D printing of tissue layer models
	<ol>
		<li>Draw cross-sectional designs of each phantom (Figure 2) and utilize computer-aided design (CAD) to design plastic models of desired tissue layers and an outer container for the overall shape of the phantom (Figure 3A, Supplemental File 1, Supplemental File 2, Supplemental File 3, and Supplemental File 4). 
		NOTE: From a design standpoint, fascial plane blocks can be thought of as a series of tightly fitted prisms enclosed within a hollow rectangular prism with 5 mm walls. Walls less than 5 mm were too fragile for reliable production. This rectangular prism serves as the outermost layer of the phantom into which the other tissue layers are assembled and placed. The median nerve model (shown in Figure 3 and Figure 4) uses an arched container rather than a rectangular prism to simulate the shape of a human arm.
		<ol>
			<li>In CAD software, create a flat canvas from an ultrasound, CT, or other target image with known dimensions. Click on Solid | Insert | Canvas, select File, click on XY plane, drag into the +X +Y space, and drag to scale such that the picture length is accurate to known units.</li>
			<li>Create a rectangular prism over the region of the image that outlines the model by clicking on Solid | Create Sketch | XY plane | 2-Point Rectangle tool. Drag the rectangle over the area and refine within the length/width boxes when close; press Enter; click on Finish Sketch; click on rectangle | Solid | Extrude; drag the rectangle to the desired height and refine with the height box when close; and press Enter. 
			NOTE: Each of our models has a different length and width based on the anatomy they represent but we typically found ~100 mm to be an effective model height.</li>
			<li>Create another sketch on top of the rectangular prism by clicking on the top of the rectangular prism | Create Sketch and draw out the desired anatomy as well as the inner edge of the encasing rectangular prism by clicking on Sketch | Create and Sketch | Modify toolboxes. Use the canvas visible behind the prism to guide the design; if the canvas is not seen through the rectangular prism, change this via Display Settings. Once the sketch that represents the cross section of the model is created, click on Finish Sketch. 
			NOTE: There is no specific ideal tool to draw the desired anatomy and the inner edge of the encasing prism. The above step is what was used in this protocol.</li>
			<li>Next, create each internal prism from the sketch by clicking on the shape in the sketch | Solid | Extrude; drag the shape back into the rectangular prism at the desired length, typically 5 mm less than the full rectangle length; and click on Operation = New Body | Enter. To view this new object, turn off the visibility of all other objects by clicking on Bodies | the eye symbol next to the new body&#39;s name. 
			NOTE: Vessels and should be represented by circular or elliptical prism-shaped holes designed into the edges or centers of fascial plane bodies. At this point, if you view the virtual model en-face to the initial canvas, the individual model pieces and how they fit together can be visualized.</li>
			<li>Export each body individually for 3D printing by clicking on Bodies | the eye symbol next to every body except the one being exported; click on File | Export | Type = .stl file | Enter. 
			NOTE: You should now have multiple .stl files, each representing a unique fascial plane or bone, as well as one additional .stl file representing the rectangular bounding box the fascial plane pieces will fit into.</li>
		</ol>
		</li>
		<li>Open the STL file in a slicer software compatible with the 3D printer that will be used to print the models.
		<ol>
			<li>Use the Place on face button to lay the model on the bed such that the bottom of the model is touching the print bed.</li>
			<li>Under Printer, select the printer. Under Print settings, select 0.20mm SPEED, and under Filament, select Generic PLA. Select 15-20% for Infill, select Everywhere on the Supports menu, and add a brim if necessary for print stability. Click Slice now.</li>
			<li>Export the G-code file to an SD card, plug it into the 3D printer, and print the file using polylactic acid (PLA) filament.</li>
		</ol>
		</li>
	</ol>
	</li>
	<li>Creation of silicone molds
	<ol>
		<li>Glue each 3D-printed model to the bottom of a topless plexiglass container before submerging it in rapid-curing silicone rubber per manufacturer guidelines17.</li>
		<li>Once the silicone is set, remove the hard-plastic model and plexiglass case, leaving a flexible, durable, and reusable silicone mold of each desired tissue layer and container into which ballistic gel is poured (Figure 3B). 
		NOTE: At this point, the protocol may be paused and restarted later.</li>
	</ol>
	</li>
</ol>
<img alt="Figure 1" class="xfigimg" src="/files/ftp_upload/66194/66194fig01.jpg" /> 
Figure 1: Representative ultrasound images obtained from a human subject. Representative images for the (A) median, (B) femoral, (C) suprainguinal fascia iliaca plane, and (D) serratus anterior plane nerve block models obtained from volunteer human subjects. Abbreviations: A = artery; V = vein; M = median nerve; F = femoral nerve; RAD = radius; U = ulna; AIIS = anterior inferior iliac spine; R =rib; SART = sartorius muscle; IL=Iliacus muscle; IO = internal oblique; SA = serratus anterior muscle; LD = latissimus dorsi muscle. <a href="https://www.jove.com/files/ftp_upload/66194/66194fig01large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 2" class="xfigimg" src="/files/ftp_upload/66194/66194fig02.jpg" /> 
Figure 2: Cross-sectional schematics of nerve block ultrasound phantoms. (A) Median, (B) femoral, (C) suprainguinal fascia iliaca plane, and (D) serratus anterior plane nerve block ultrasound phantoms. Schematics were designed based on the representative human ultrasound images shown in Figure 1. <a href="https://www.jove.com/files/ftp_upload/66194/66194fig02large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 3" class="xfigimg" src="/files/ftp_upload/66194/66194fig03.jpg" /> 
Figure 3: Creation of median nerve block phantom components. (A) Representative image of the computer-aided design file used to print plastic models of each tissue layer for the median nerve block phantom. (B) Silicone molds for each tissue layer of the median nerve block phantom, including metal rods inserted to create vessels within the ballistic gel. (C) Pouring hot, liquid, dyed ballistic gel into the silicone molds. (D) Sealing the open end of simulated vessels using liquid ballistic gel after vessels have been filled with simulated blood. <a href="https://www.jove.com/files/ftp_upload/66194/66194fig03large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
2. Creation of other phantom landmarks
<ol>
	<li>Simulated bone design and creation
	<ol>
		<li>If the region of the model designed in CAD represents bone instead of soft tissue, 3D print an imitation bone using the steps above, but use acrylonitrile butadiene styrene (ABS) filament instead. 
		CAUTION: ABS filament fumes may contain volatile organic compounds (VOCs), which can cause physical discomfort, such as drowsiness, eye or respiratory tract irritation, nausea, and/or headaches18. ABS should be printed on an enclosed 3D printer or an unenclosed printer in a room with good ventilation and/or air filtration.</li>
	</ol>
	</li>
	<li>Simulated nerve creation
	<ol>
		<li>Submerge 80% acrylic, 20% wool yarn in a plastic cup filled with ultrasound gel. Place the cup is placed in a -1 atm pressure chamber.</li>
		<li>Use a single-stage vacuum pump to repeatedly build up and then release pressure in the chamber until all bubbles have been removed from the ultrasound gel, accomplished after approximately 4-6 cycles. 
		&#8203;NOTE: This step helps simulate nerves. At this point, the protocol may be paused and restarted later.</li>
	</ol>
	</li>
</ol>
3. Ballistic gel melting and pouring
<ol>
	<li>Melting the ballistic gel
	<ol>
		<li>Heat commercially available ballistic gel and dye at an approximate 20:1 volume ratio with intermittent stirring until the liquid reaches 132 &#176;C in a commercial convection oven. 
		CAUTION: Heated, liquid ballistic gel should be handled with caution due to the risk of burns associated with handling hot fluids. Use oven mitts when handling pans filled with liquid ballistic gel. Avoid direct contact between skin and liquid ballistic gel.</li>
	</ol>
	</li>
	<li>Additives for echogenicity
	<ol>
		<li>Stir approximately 4.5 g of finely granulated flour per kg of ballistic gel into the liquid ballistic gel. Leave the gel in the oven for at least 20 min with intermittent stirring to allow for even mixing and to allow any bubbles to escape.</li>
		<li>Add additional clear ballistic gel or dye as necessary to adjust the mixture&#39;s color to simulate human tissue.</li>
	</ol>
	</li>
	<li>Ballistic gel pouring into silicone molds
	<ol>
		<li>Insert solid steel rods of varying diameters into the designated locations on the reusable silicone molds, if appropriate for that specific nerve block model, to create channels in the final ultrasound phantoms, which will represent blood vessels (Figure 3C).</li>
		<li>Pour liquid ballistic gel, now colored by dye, with suspended flour particulates, and without retained air bubbles, into the silicone molds and allow it to cool.</li>
		<li>After cooling, remove the metal rods and the final ballistic gel tissue layers from the molds. When placed together with an ultrasound gel coating between them, adjacent tissue pieces align near-perfectly and together produce a simulated fascial plane on ultrasound. 
		NOTE: Each ultrasound phantom requires approximately 0.7 kg of ballistic gel. Cooling time is dependent on the size of the tissue layer and varies from 20 min to 1.5 h.</li>
	</ol>
	</li>
	<li>Addition of simulated blood and sealing of vessels
	<ol>
		<li>For the tissue layers with simulated vessels, dip one side of the tissue layer into the liquid ballistic gel and allow it to cool, thus sealing off one side of the vessel channel.</li>
		<li>Hold these tissue layers upright and use a needle and syringe to introduce simulated blood into each vessel. 
		NOTE: We used water with red or blue food coloring to represent arterial and venous blood, respectively.</li>
		<li>Use still-liquid ballistic gel to cover up the remaining vessel opening, thus sealing off each fluid-filled vessel completely (Figure 3D). 
		&#8203;NOTE: At this point, the protocol may be paused and restarted later; however, ballistic gel must be melted again to proceed to the following step.</li>
	</ol>
	</li>
</ol>
4. Phantom assembly
<ol>
	<li>Assembly of tissue layers, nerves, and bones 
	NOTE: Figure 4A depicts the individual components of the median nerve block phantom immediately prior to assembly, including the tissue layers, simulated nerves, and simulated bones.
	<ol>
		<li>Assemble the phantoms by coating each component in ultrasound gel, assembling the components as shown by the cross-sections in Figure 2, and inserting them into their respective ballistic gel rectangular prisms (Figure 4B). Place any 3D-printed bones or yarn nerves properly at this step.</li>
	</ol>
	</li>
	<li>Sealing the ends of the phantom
	<ol>
		<li>Seal the models by dipping them on both sides into a pan filled with liquid ballistic gel (Figure 4C). Repeat the sealing process multiple times on each side.</li>
		<li>Finally, use a heat gun to smooth the edges of the phantom, removing bubbles and imperfections, as well as reinforcing the side seals.</li>
	</ol>
	</li>
	<li>Addition of pseudo-skin (optional) 
	NOTE: An augmentation to the fascial-plane models is the addition of pseudo-skin.
	<ol>
		<li>Pour liquid ballistics gel over a sealed and cooled model, which has been covered loosely in ultrasound gel to prevent annealing between the newly poured skin layer and the existing model (Figure 4D and Supplemental Video S1).</li>
	</ol>
	</li>
</ol>
<img alt="Figure 4" class="xfigimg" src="/files/ftp_upload/66194/66194fig04.jpg" /> 
Figure 4: Assembly of median nerve block ultrasound phantom. (A) Individual components of a disassembled median nerve block phantom, including ballistic gel tissue layers, 3D-printed radius and ulna, a yarn median nerve submerged in ultrasound gel, a bottle of ultrasound gel, and a pan filled with liquid ballistic gel. (B) Assembly of the median nerve block phantom, including insertion of tissue layers and simulated bones covered in ultrasound gel. (C) Sealing one end of the phantom by dipping into a pan of liquid ballistic gel. (D) Creating a layer of pseudo-skin by pouring liquid ballistic gel over a completed median nerve block phantom. <a href="https://www.jove.com/files/ftp_upload/66194/66194fig04large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>

Building Custom Ballistic Gel Ultrasound Phantoms for Nerve Block Simulations

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The authors of this paper do not have any conflicts of interest to disclose.

These custom ballistic gel-based phantoms provide trainees with medium-fidelity median, femoral, suprainguinal fascia iliaca plane, and serratus anterior plane nerve block training for a fraction of the cost of commercially available nerve block phantoms (Table 1). Our first median and femoral nerve block trainers were made in-house for 12% and 9% of the price of the cheapest commercially available median and femoral nerve block trainers, respectively. None of the available femoral nerve block phantoms a...

Ultrasound phantoms - alternatives to live human tissue - give learners the opportunity to practice medical procedures, including ultrasound-guided regional anesthesia (UGRA), without introducing undue risk to patients1. While most commonly manufactured via injection molding of liquid silicone rubber, custom phantoms can be homemade using versatile materials at lower cost. Organic tissues such as tofu, pork, and beef are inexpensive but spoil quickly and are challenging to craft2. Human cadaveric tissue is ideal for anatomic accuracy but is difficult and costly to obtain and preserve1. More recently, virtual reality has been used to provide UGRA training; however, haptic feedback is a key component of procedural learning and is rarely implemented. Even when a hardware-software hybrid model provides high visual fidelity and tactile feedback, the hardware and software required to carry out such training are frequently cost-prohibitive3. Gelatin-based phantoms strike a balance between cost, longevity, and fidelity2.
Ballistic gelatin models are available commercially but are expensive for a perishable resource that is highly utilized in medical simulation centers. Small, simple, gel-based ultrasound phantoms with homogeneous parenchyma and two or three simulated vessels retail for hundreds of dollars. For example, the CAE Blue Phantom basic ultrasound training block costs upwards of $8004. Higher-fidelity phantoms specific to individual nerve block procedures cost thousands of dollars. The CAE Blue Phantom femoral regional anesthesia ultrasound training model costs $5,000 (Table 1)5. To bring down costs, educators have experimented with custom-made phantoms using gelatin or other low-cost or reusable materials6,7,8. Additives such as flour, corn starch, graphite powder, and Metamucil can be used to opacify the gelatin and customize the echogenicity of the phantom, thus increasing its fidelity8,9,10,11,12,13,14.
Previous attempts at homemade gelatin-based nerve block trainers were either unable to adequately recreate the appearance of nerves under ultrasound or utilized perishable items, thus limiting shelf life15,16. Even without these drawbacks, previous iterations did not include relevant anatomical landmarks and fascial planes that would allow trainees to practice specific nerve block procedures. Here, we investigate the production of durable, low-cost, ballistic gel ultrasound phantoms for median, femoral, suprainguinal fascia iliaca plane, and serratus anterior plane nerve blocks, as well as a methodology for producing a phantom for any ultrasound-guided nerve block procedure.

<table><tbody><tr><td>ABS Filament - 1.75 m+B+A2:A14</td><td>Hatchbox (Pomona, CA)</td><td></td><td>https://www.hatchbox3d.com/collections/abs-1-75mm</td></tr><tr><td>Adobe Photoshop</td><td>Adobe (San Jose, CA)</td><td></td><td>https://www.adobe.com/products/photoshop.html</td></tr><tr><td>Amber Tone Dye</td><td>Humimic Medical (Greenville, SC)</td><td>852844007925</td><td>Ballistic gel dye; https://humimic.com/product/amber-tone-dye/</td></tr><tr><td>Fusion 360</td><td>Autodesk (San Franciso, CA)</td><td></td><td>Computer-assisted design (CAD) software; https://www.autodesk.com/products/fusion-360/overview?term=1-YEAR&amp;amp;tab=subscription&amp;amp;plc=F360</td></tr><tr><td>Gelatin #1 - Medical Gel by the Pound</td><td>Humimic Medical (Greenville, SC)</td><td>852844007406</td><td>Ballistic gel; https://humimic.com/product/gelatin-1-medical-gelatin-by-the-pound/</td></tr><tr><td>Gluten-Free All-Purpose Flour</td><td>Arrowhead Mills (Hereford, TX)</td><td></td><td>Flour for echogenicity; https://arrowheadmills.com/products/gluten-free/organic-gluten-free-all-purpose-flour/</td></tr><tr><td>Microsoft PowerPoint</td><td>Microsoft (Redmond, WA)</td><td></td><td>https://www.microsoft.com/en-us/microsoft-365/powerpoint</td></tr><tr><td>Mold Star 16 FAST Pourable Silicone Rubber</td><td>Smooth-On (Macungie, PA)</td><td></td><td>Pourable silicone rubber; https://www.smooth-on.com/products/mold-star-16-fast/</td></tr><tr><td>Peach Tone Dye</td><td>Humimic Medical (Greenville, SC)</td><td>852844007895</td><td>Ballistic gel dye; https://humimic.com/product/peach-tone-dye/</td></tr><tr><td>PLA Filament - 1.75 mm</td><td>Hatchbox (Pomona, CA)</td><td></td><td>https://www.hatchbox3d.com/collections/pla-1-75mm</td></tr><tr><td>Prusa Original i3 MK3S+ printer</td><td>Prusa Research (Prague, Czech Republic)</td><td></td><td>3D printer; https://www.prusa3d.com/category/original-prusa-i3-mk3s/</td></tr><tr><td>Prusa Slicer 2.6.1</td><td>Prusa Research (Prague, Czech Republic)</td><td></td><td>3D printer slicer software; https://www.prusa3d.com/page/prusaslicer_424/</td></tr><tr><td>Wool-Ease Thick &amp;amp; Quick Yarn</td><td>Lion Brand Yarn (Lyndhurst, NJ)</td><td>640-610B</td><td>Yarn for simulated nerves; https://www.lionbrand.com/products/wool-ease-thick-and-quick-yarn?variant=32420963516509</td></tr></tbody></table>

building affordable, durable, medium-fidelity ballistic gel phantoms for ultrasound-guided nerve block training

Ultrasound phantoms - alternatives to live human tissue - give learners the opportunity to practice ultrasound-guided regional anesthesia without introducing undue risk to patients. Gelatin-based phantoms provide educators with durable and reusable task trainers; however, commercially available gel-based phantoms are expensive. Here, we investigate the production of durable, low-cost, ballistic gel-based ultrasound phantoms for median, femoral, suprainguinal fascia iliaca plane, and serratus anterior plane nerve blocks, as well as a methodology for producing a phantom for any ultrasound-guided nerve block procedure.
Computer-aided design (CAD) software was utilized to design four phantoms replicating the anatomy of median, femoral, suprainguinal fascia iliaca plane, and serratus anterior plane nerve blocks, including relevant landmarks and tissue planes. Plastic models of the desired tissue planes were 3D printed and used to create silicone molds. Ballistic gel was melted and mixed with flour and dye to create a liquid, echogenic ballistic gel, which was poured into the silicone molds. Vessels were simulated by creating negative space in the ballistic gel using metal rods. Nerves were simulated using yarn submerged in ultrasound gel. Simulated bones were designed using CAD and 3D printed.
Ballistic gel is a versatile, durable medium that can be used to simulate a variety of tissues and can be melted and molded into any shape. Under ultrasound, these phantoms provide realistic tissue planes that represent the borders between different layers of skin, muscle, and fascia. The echogenicity of the muscle tissue layers, nerves, vessels, and bones is realistic, and bones have significant posterior shadowing as would be observed in a human subject. These phantoms cost $200 each for the first phantom and $60 for each subsequent phantom. These phantoms require some technical skill to design, but they can be built for just 4% of the cost of their commercial counterparts.

Four ultrasound phantoms were successfully designed and built using the methods described above. An ultrasound cross-section of each model aligned with an ultrasound of equivalent human anatomy is shown in Figure 5. Under ultrasound, these phantoms provide realistic tissue planes, which represent the borders between different layers of skin, muscle, and fascia. The muscle tissue is appropriately and homogeneously echogenic. This echogenicity can be adjusted based on the amount of flour added...

Read this Scientific Article Protocol about Author Spotlight: Developing Cost-Effective and Durable Ultrasound-Guided 3D-Printed Nerve Block Trainers at JoVE.com

Author Spotlight: Developing Cost-Effective and Durable Ultrasound-Guided 3D-Printed Nerve Block Trainers

Here, we demonstrate the design and creation of four custom ballistic gelatin ultrasound phantoms for ultrasound-guided regional anesthesia training. We designed the phantoms using computer-aided design software, used 3D-printed models to create silicone molds, and then poured melted ballistic gel into the molds to create custom tissue layers.

Building Affordable, Durable, Medium-Fidelity Ballistic Gel Phantoms for Ultrasound-Guided Nerve Block Training

Ultrasound phantoms - alternatives to live human tissue - give learners the opportunity to practice ultrasound-guided regional anesthesia without ...

building-affordable-durable-medium-fidelity-ballistic-gel-phantoms

Research

JoVE Journal

Medicine

1.6K Views.  University of California, San Diego School of Medicine. Daarom hebben we gebruik gemaakt van de nieuwste technologie, zoals computerondersteund ontwerp en 3D-printen, om op maat gemaakte, duurzame, echogeleide zenuwblokkadetrainers te maken met relatief goedkope materialen. Daarbij onderzochten we vragen met betrekking tot de duurzaamheid en kosteneffectiviteit van zenuwblokkadetrainers, terwijl we de betrouwbaarheid en toegankelijkheid van deze trainers maximaliseerden. Momenteel moeten medische opvoeders en simulatiecentra ofwel duizenden dollar...

Video: Auteur in de schijnwerpers: ontwikkeling van kosteneffectieve en duurzame ultrageleide 3D-geprinte zenuwblokbloktrainers

Controllable Nucleation of Cavitation from Plasmonic Gold Nanoparticles for Enhancing High Intensity Focused Ultrasound Applications

In this study, plasmonic gold nanoparticles were simultaneously exposed to pulsed near-infrared laser light and high intensity focused ultrasound (HIFU) for the controllable nucleation of cavitation in tissue-mimicking gel phantoms. This in vitro protocol was developed to demonstrate the feasibility of this approach, for both enhancement of imaging and therapeutic applications for cancer. The same apparatus can be used for both imaging and therapeutic applications by varying the exposure duration of the HIFU system. For short duration exposures (10 &#181;s), broadband acoustic emissions were generated through the controlled nucleation of inertial cavitation around the gold nanoparticles. These emissions provide direct localization of nanoparticles. For future applications, these particles may be functionalized with molecular-targeting antibodies (e.g. anti-HER2 for breast cancer) and can provide precise localization of cancerous regions, complementing routine diagnostic ultrasound imaging. For continuous wave (CW) exposures, the cavitation activity was used to increase the localized heating from the HIFU exposures resulting in larger thermal damage in the gel phantoms. The acoustic emissions generated from inertial cavitation activity during these CW exposures was monitored using a passive cavitation detection (PCD) system to provide feedback of cavitation activity. Increased localized heating was only achieved through the unique combination of nanoparticles, laser light and HIFU. Further validation of this technique in pre-clinical models of cancer is necessary.

This protocol demonstrates the controllable nucleation of cavitation in gel phantoms, through simultaneous exposure to both near-infrared pulsed laser light and high intensity focused ultrasound (HIFU). The cavitation activity can then be used for enhancing imaging and/or therapeutic uses of HIFU.

In this study, plasmonic gold nanoparticles were simultaneously exposed to pulsed near-infrared laser light and high intensity focused ultrasound (HIFU) ...

Engineering

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 ...

An Ultrasonic Tool for Nerve Conduction Block in Diabetic Rat Models

Nerve conduction block with a high intensity-focused ultrasound (HIFU) transducer has been performed in normal and diabetic animal models recently. HIFU can reversibly block the conduction of peripheral nerves without damaging the nerves while using an appropriate ultrasonic parameter. Temporary and partial block of the action potentials of nerves shows that HIFU has the potential to be a useful clinical treatment for pain relief. This work demonstrates the procedures for suppressing the action potentials of neuropathic nerves in diabetic rats in vivo using an HIFU transducer. The first step is to generate adult male diabetic neuropathic rats by streptozotocin (STZ) injection. The second step is to evaluate the peripheral diabetic neuropathy in STZ-induced diabetic rats by an electronic von Frey probe and a hot plate. The final step is to record in vivo extracellular action potentials of the nerve exposed to HIFU sonication. The method showed here may benefit the study of ultrasound analgesic applications.

This work presents the methodology of applying high intensity-focused ultrasound to block the action potentials of diabetic neuropathic nerves.

Nerve conduction block with a high intensity-focused ultrasound (HIFU) transducer has been performed in normal and diabetic animal models recently. HIFU ...

Bioengineering

An Experimental Protocol for Assessing the Performance of New Ultrasound Probes Based on CMUT Technology in Application to Brain Imaging

The possibility to perform an early and repeatable assessment of imaging performance is fundamental in the design and development process of new ultrasound (US) probes. Particularly, a more realistic analysis with application-specific imaging targets can be extremely valuable to assess the expected performance of US probes in their potential clinical field of application.
The experimental protocol presented in this work was purposely designed to provide an application-specific assessment procedure for newly-developed US probe prototypes based on Capacitive Micromachined Ultrasonic Transducer (CMUT) technology in relation to brain imaging.
The protocol combines the use of a bovine brain fixed in formalin as the imaging target, which ensures both realism and repeatability of the described procedures, and of neuronavigation techniques borrowed from neurosurgery. The US probe is in fact connected to a motion tracking system which acquires position data and enables the superposition of US images to reference Magnetic Resonance (MR) images of the brain. This provides a means for human experts to perform a visual qualitative assessment of the US probe imaging performance and to compare acquisitions made with different probes. Moreover, the protocol relies on the use of a complete and open research and development system for US image acquisition, i.e. the Ultrasound Advanced Open Platform (ULA-OP) scanner.
The manuscript describes in detail the instruments and procedures involved in the protocol, in particular for&#160;the calibration, image acquisition and registration of US and MR images. The obtained results prove the effectiveness of the overall protocol presented, which is entirely open (within the limits of the instrumentation involved), repeatable, and covers the entire set of acquisition and processing activities for US images.

The development of new ultrasound (US) probes based on Capacitive Micromachined Ultrasonic Transducer (CMUT) technology requires an early realistic assessment of imaging capabilities. We describe a repeatable experimental protocol for US image acquisition and comparison with magnetic resonance images, using an ex vivo bovine brain as an imaging target.

The possibility to perform an early and repeatable assessment of imaging performance is fundamental in the design and development process of new ...

Patient-Specific Polyvinyl Alcohol Phantom Fabrication with Ultrasound and X-Ray Contrast for Brain Tumor Surgery Planning

Phantoms are essential tools for clinical training, surgical planning and the development of novel medical devices. However, it is challenging to create anatomically accurate head phantoms with realistic brain imaging properties because standard fabrication methods are not optimized to replicate any patient-specific anatomical detail and 3D printing materials are not optimized for imaging properties. In order to test and validate a novel navigation system for use during brain tumor surgery, an anatomically accurate phantom with realistic imaging and mechanical properties was required. Therefore, a phantom was developed using real patient data as input and 3D printing of molds to fabricate a patient-specific head phantom comprising the skull, brain and tumor with both ultrasound and X-ray contrast. The phantom also had mechanical properties that allowed the phantom tissue to be manipulated in a similar manner to how human brain tissue is handled during surgery. The phantom was successfully tested during a surgical simulation in a virtual operating room.
The phantom fabrication method uses commercially available materials and is easy to reproduce. The 3D printing files can be readily shared, and the technique can be adapted to encompass many different types of tumor.

This protocol describes the fabrication of a patient specific skull, brain and tumor phantom. It uses 3D printing to create molds, and polyvinyl alcohol (PVA-c) is used as the tissue mimicking material.

Phantoms are essential tools for clinical training, surgical planning and the development of novel medical devices. However, it is challenging to create ...

Viscoelastic Characterization of Soft Tissue-Mimicking Gelatin Phantoms using Indentation and Magnetic Resonance Elastography

Characterization of biomechanical properties of soft biological tissues is important to understand the tissue mechanics and explore the biomechanics-related mechanisms of disease, injury, and development. The mechanical testing method is the most straightforward way for tissue characterization and is considered as verification for in vivo measurement. Among the many ex vivo mechanical testing techniques, the indentation test provides a reliable way, especially for samples that are small, hard to fix, and viscoelastic such as brain tissue. Magnetic resonance elastography (MRE) is a clinically used method to measure the biomechanical properties of soft tissues. Based on shear wave propagation in soft tissues recorded using MRE, viscoelastic properties of soft tissues can be estimated in vivo based on wave equation. Here, the viscoelastic properties of gelatin phantoms with two different concentrations were measured by MRE and indentation. The protocols of phantom fabrication, testing, and modulus estimation have been presented.

This article presents a demonstration and summary of protocols of making gelatin phantoms that mimic soft tissues, and the corresponding viscoelastic characterization using indentation and magnetic resonance elastography.

Characterization of biomechanical properties of soft biological tissues is important to understand the tissue mechanics and explore the ...

A High-Throughput Image-Guided Stereotactic Neuronavigation and Focused Ultrasound System for Blood-Brain Barrier Opening in Rodents

The blood-brain barrier (BBB) has been a major hurdle for the treatment of various brain diseases. Endothelial cells, connected by tight junctions, form a physiological barrier preventing large molecules (&#62;500 Da) from entering the brain tissue. Microbubble-mediated focused ultrasound (FUS) can be used to induce a transient local BBB opening, allowing larger drugs to enter the brain parenchyma.
In addition to large-scale clinical devices for clinical translation, preclinical research for therapy response assessment of drug candidates requires dedicated small animal ultrasound setups for targeted BBB opening. Preferably, these systems allow high-throughput workflows with both high-spatial precision as well as integrated cavitation monitoring, while still being cost effective in both initial investment and running costs.
Here, we present a bioluminescence and X-ray guided stereotactic small animal FUS system that is based on commercially available components and fulfills the aforementioned requirements. A particular emphasis has been placed on a high degree of automation facilitating the challenges typically encountered in high-volume preclinical drug evaluation studies. Examples of these challenges are the need for standardization in order to ensure data reproducibility, reduce intra-group variability, reduce sample size and thus comply with ethical requirements and decrease unnecessary workload. The proposed BBB system has been validated in the scope of BBB opening facilitated drug delivery trials on patient-derived xenograft models of glioblastoma multiforme and diffuse midline glioma.

The blood-brain barrier (BBB) can be temporarily disrupted with microbubble-mediated focused ultrasound (FUS). Here, we describe a step-by-step protocol for high-throughput BBB opening in vivo using a modular FUS system accessible for non-ultrasound experts.

The blood-brain barrier (BBB) has been a major hurdle for the treatment of various brain diseases. Endothelial cells, connected by tight junctions, form a ...

Neuroscience

Focused Ultrasound Induced Blood-Brain Barrier Opening for Targeting Brain Structures and Evaluating Chemogenetic Neuromodulation

Acoustically Targeted Chemogenetics (ATAC) allows for the noninvasive control of specific neural circuits. ATAC achieves such control through a combination of focused ultrasound (FUS) induced blood-brain barrier opening (FUS-BBBO), gene delivery with adeno-associated viral (AAV) vectors, and activation of cellular signaling with engineered, chemogenetic, protein receptors and their cognate ligands. With ATAC, it is possible to transduce both large and small brain regions with millimeter precision using a single noninvasive ultrasound application. This transduction can later allow for a long-term, noninvasive, device-free neuromodulation in freely moving animals using a drug. Since FUS-BBBO, AAVs, and chemogenetics have been used in multiple animals, ATAC should also be scalable for the use in other animal species. This paper expands upon a previously published protocol and outlines how to optimize the gene delivery with FUS-BBBO to small brain regions with MRI-guidance but without a need for a complicated MRI-compatible FUS device. The protocol, also, describes the design of mouse targeting and restraint components that can be 3D-printed by any lab and can be easily modified for different species or custom equipment. To aid reproducibility, the protocol describes in detail how the microbubbles, AAVs, and venipuncture were used in ATAC development. Finally, an example data is shown to guide the preliminary investigations of studies utilizing ATAC.

This protocol delineates steps necessary for the gene delivery through focused ultrasound blood brain barrier (BBB) opening, evaluation of the resulting gene expression, and measurement of neuromodulation activity of chemogenetic receptors through histological tests.

Acoustically Targeted Chemogenetics (ATAC) allows for the noninvasive control of specific neural circuits. ATAC achieves such control through a ...

High-frequency Ultrasonography of the Digital and Palmar Nerve Branches of the Hand in Peripheral Nerve Diseases

Peripheral nerve ultrasound is a well-established imaging technique to evaluate certain peripheral nerve pathologies. However, there is a poor correlation between ultrasound abnormalities of peripheral nerves and electrodiagnostic or clinical evidence of axonal loss. This is a significant limitation of peripheral nerve ultrasound, as many peripheral nerve diseases encountered in clinical settings are related to axonal loss. Furthermore, clinical and electrodiagnostic evidence of axonal loss directly correlates with disability in all peripheral nerve diseases. However, due to the floor effects often encountered in electrodiagnostic studies, these correlations, as well as definitive diagnoses, are often challenging. Thus, imaging techniques that correlate with axonal loss are essential for expanding the utility of peripheral nerve ultrasound as a potential biomarker for peripheral nerve diseases. With new technological advancements and the ever-increasing imaging capabilities of high-frequency ultrasound, the palmar and digital nerve branches of the hand can be imaged with exceptionally high resolution even using point-of-care ultrasound devices. Their superficial and distal-most anatomic locations are ideal for evaluating polyneuropathies, as these branches degenerate earliest during axonal loss. However, no studies have systematically evaluated these nerve branches to determine if they can be reproducibly measured with ultrasound. The current protocol was adapted for the systematic assessment of cross-sectional areas of the median and ulnar nerves in the palmar surface and digits of the hand. This protocol provides reference data for a subset of nerves that demonstrate high intraclass correlation coefficients between three separate ultrasonographers. Finally, as a proof of concept and to demonstrate the clinical applications of this protocol, representative data from individuals with genetically confirmed inherited polyneuropathies are compared with established normative data to examine cross-sectional area differences.

The present protocol describes high-frequency neuromuscular ultrasonography of the digital and palmar branches of the median and ulnar nerve, which can aid in localizing peripheral nerve diseases and be adapted to evaluate digital nerve injuries.

Peripheral nerve ultrasound is a well-established imaging technique to evaluate certain peripheral nerve pathologies. However, there is a poor correlation ...

Nerve Ultrasound Protocol to Detect Dysimmune Neuropathies

Nerve ultrasound is increasingly used in the differential diagnosis of polyneuropathy as a complementary tool to nerve conduction studies. Morphological alterations of the peripheral nerves, such as increasing the cross-sectional area (CSA), have been described in various immune-mediated polyneuropathies. The most prominent morphological changes in nerve ultrasound have been described for the chronic inflammatory demyelinating polyneuropathy (CIDP)-spectrum disease. CIDP may be distinguished from hereditary and other polyneuropathies by measuring the extent and pattern of nerve swellings (CSA increase). Typical findings in demyelinating inflammatory neuropathies are multifocal nerve swellings with inhomogeneous fascicular structure, while CSA increase in demyelinating hereditary neuropathies occurs in a more generalized and homogenous manner. In other non-inflammatory axonal neuropathies, nerves can appear with normal or slight CSA increases, especially in typical entrapment sites. This article presents technical requirements for nerve ultrasound, an examination procedure using a standardized examination protocol, current reference values for the CSA, and typical sonographic pathological findings in patients with inflammatory neuropathies.

This article presents a protocol for nerve ultrasound in polyneuropathies to aid the diagnosis of inflammatory neuropathies.

Nerve ultrasound is increasingly used in the differential diagnosis of polyneuropathy as a complementary tool to nerve conduction studies. Morphological ...

Building Affordable, Durable, Medium-Fidelity Ballistic Gel Phantoms for Ultrasound-Guided Nerve Block Training

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