Name: fat-water phantoms for magnetic resonance imaging validation: a flexible and scalable protocol
Uploaded: 2018-09-07T13:45:00
Description: Watch this Scientific Journal Video about Fat-Water Phantoms for Magnetic Resonance Imaging Validation: A Flexible and Scalable Protocol at JoVE.com

Funding support for this research was provided the National Institutes of Health (NIH) and National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)/NIH R01-DK-105371. We thank Dr. Houchun (Harry) Hu for advice and suggestions on fat water phantom creation.

1. Prepare the Workstation and Materials
<ol>
	<li>Adhere to all laboratory safety rules. Wear eye protection and gloves. Read the material safety data sheet for each of the reagents used and take appropriate precautions. Review the materials and equipment list, chemical handling procedures, and glassware precautions. 
	Caution: This protocol requires the use of a hotplate at high temperatures. Use caution and wear heat resistant gloves when interacting with hot containers and do not touch the surface of the hotpl...

<ol>
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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=Liver+Fat+Quantification+by+Dual-echo+MR+Imaging+Outperforms+Traditional+Histopathological+Analysis.">Liver Fat Quantification by Dual-echo MR Imaging Outperforms Traditional Histopathological Analysis.</a> Acad Radiol. 19 (10), 1208-1214 (2012).</li><li>Hayashi, T., 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=Influence+of+Gd-EOB-DTPA+on+proton+density+fat+fraction+using+the+six-echo+Dixon+method+in+3+Tesla+magnetic+resonance+imaging.">Influence of Gd-EOB-DTPA on proton density fat fraction using the six-echo Dixon method in 3 Tesla magnetic resonance imaging.</a> Radiol Phys Technol. , (2017).</li><li>Hines, C. D. G., Yu, H., Shimakawa, A., McKenzie, C. A., Brittain, J. H., Reeder, S. 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We describe a robust method to create fat water phantoms suitable for validating the medical imaging techniques used to quantify adipose tissue and triglyceride content in vivo. By creating two reservoirs (one for the oil solution and one for the water solution), stable phantoms with a variety of FF values &#8211; including values exceeding 50% &#8211; were constructed without the need for expensive equipment. High FF phantoms (&#62;50%) provide the utility to ensure imaging techniques for adipose quantification...

Interest in quantifying adipose tissue and triglyceride content using imaging modalities, such as magnetic resonance imaging (MRI), extends across many fields. Research areas include the investigation of white and brown adipose tissue depots and ectopic storage of lipid in organs and tissues such as the liver1, pancreas2, and skeletal muscle3. As these novel techniques for adipose quantification are developed, methods are needed to confirm that imaging parameters are valid for research and clinical applications.
Phantoms, experimental replicas of a tissue or organ, provide a low-cost, flexible, and controlled tool to develop and validate imaging techniques4. Specifically, phantoms can be constructed to consist of fat and water in a volume ratio or fat fraction (FF) comparable to that of the tissue of clinical interest. Clinically, FF values in tissues and organs can vary widely: FF in brown adipose tissue falls between 29.7% and 93.9%5; the average liver FF in steatosis patients is 18.1 &#177; 9.0%6; the pancreatic FF in adults at risk for type 2 diabetes ranges between 1.6% and 22.2%7; and in some cases of advance disease, patients with Duchenne muscular dystrophy can have FF values of almost 90% in some muscles8.
Because non-polar molecules such as lipids do not dissolve well in solutions composed of polar molecules such as water, creating stable phantoms with a high target FF remains challenging. For FF up to 50%, many existing methods can be used to create fat water phantoms9,10,11,12. Other methods that achieve higher FFs typically require expensive equipment such as a homogenizer or an ultrasonic cell disruptor13,14. Although these techniques provide a roadmap for high FF phantoms, equipment constraints and varying amounts of experimental details limit the efforts to create reproducible and robust fat water phantoms.
Building upon these previous techniques, we developed a method to construct cost-effective and stable fat water phantoms across a customizable range of FF values. This protocol details the steps needed to make 5x 100 mL of fat phantoms with FF values of 0%, 25%, 50%, 75%, and 100% using a single hotplate. It can easily be adjusted to create various volumes (10 to 200 mL) and fat percentages (0 to 100%). The efficacy of the phantom technique was evaluated in the feasibility study comparing fat-water MRI FF values to the target FF values in the constructed phantoms.

<table>
	<tbody>
		<tr>
			<td>Distilled Water</td>
			<td>Amazon</td>
			<td>B000P9BY38</td>
			<td>Base of water solution</td>
		</tr>
		<tr>
			<td>Agar</td>
			<td>Sigma Aldrich Incorporated</td>
			<td>A1296-100G</td>
			<td>Gelling agent</td>
		</tr>
		<tr>
			<td>Water-Soluble Surfactant</td>
			<td>Sigma Aldrich Incorporated</td>
			<td>P1379-500ML</td>
			<td>Surfactant/emulsifying agent</td>
		</tr>
		<tr>
			<td>Gadolinium-DTPA Contrast Agent</td>
			<td>Bayer Healthcare</td>
			<td>50419-0188-01</td>
			<td>Magnetic Resonance Imaging Contrast Agent.</td>
		</tr>
		<tr>
			<td>Sodium Benzoate</td>
			<td>Sigma Aldrich Incorporated</td>
			<td>71300-250G</td>
			<td>Preservative</td>
		</tr>
		<tr>
			<td>Peanut Oil</td>
			<td>Amazon</td>
			<td>54782-LOU</td>
			<td>Base of oil solution</td>
		</tr>
		<tr>
			<td>Oil-Soluble Surfactant</td>
			<td>Sigma Aldrich Incorporated</td>
			<td>S6760-250ML</td>
			<td>Surfactant/emulsifying agent</td>
		</tr>
		<tr>
			<td>Hotplate w/ Stirrer</td>
			<td>Fisher Scientific</td>
			<td>07-770-152</td>
			<td></td>
		</tr>
		<tr>
			<td>Stir bars (Egg-Shaped)</td>
			<td>Sigma Aldrich Incorporated</td>
			<td>Z127116-1EA</td>
			<td></td>
		</tr>
		<tr>
			<td>400 mL Beaker</td>
			<td>Sigma Aldrich Incorporated</td>
			<td>CLS1003400-48EA</td>
			<td></td>
		</tr>
		<tr>
			<td>250 mL Erlenmeyer Flask</td>
			<td>Sigma Aldrich Incorporated</td>
			<td>CLS4450250-6EA</td>
			<td></td>
		</tr>
		<tr>
			<td>25 mL Glass Volumetric Pipette</td>
			<td>Fisher Scientific</td>
			<td>13-650-2P</td>
			<td>Quantity = 2</td>
		</tr>
		<tr>
			<td>50 mL Glass Volumetric Pipette</td>
			<td>Fisher Scientific</td>
			<td>13-650-2S</td>
			<td>Quantity = 2</td>
		</tr>
		<tr>
			<td>75 mL Glass Volumetric Pipette</td>
			<td>Fisher Scientific</td>
			<td>13-650-2T</td>
			<td>Quantity = 2</td>
		</tr>
		<tr>
			<td>3.0 mL Syringe</td>
			<td>Sigma Aldrich Incorporated</td>
			<td>Z248002-1PAK</td>
			<td></td>
		</tr>
		<tr>
			<td>1.0 mL Syringe</td>
			<td>Sigma Aldrich Incorporated</td>
			<td>Z230723-1PAK</td>
			<td></td>
		</tr>
		<tr>
			<td>Spatula</td>
			<td>Sigma Aldrich Incorporated</td>
			<td>S3897-1EA</td>
			<td></td>
		</tr>
		<tr>
			<td>Scale (100g X 0.01g Resolution)</td>
			<td>Amazon</td>
			<td>AWS-100-BLK</td>
			<td></td>
		</tr>
		<tr>
			<td>Weigh Boats</td>
			<td>Sigma Aldrich Incorporated</td>
			<td>Z740499-500EA</td>
			<td></td>
		</tr>
		<tr>
			<td>120 mL Glass Jars</td>
			<td>McMaster Carr Supply Co</td>
			<td>3801T73</td>
			<td></td>
		</tr>
		<tr>
			<td>Heat Resistant Gloves (pair)</td>
			<td>Amazon</td>
			<td>B075GX43MN</td>
			<td></td>
		</tr>
		<tr>
			<td>Syringe Needles</td>
			<td>Sigma Aldrich Incorporated</td>
			<td>Z192341-100EA</td>
			<td></td>
		</tr>
		<tr>
			<td>18&#34; stir bar retriver</td>
			<td>Fisher Scientific</td>
			<td>14-513-70</td>
			<td></td>
		</tr>
		<tr>
			<td>1 Dram Clear Glass Vial</td>
			<td>Fisher Scientific</td>
			<td>03-339-25B</td>
			<td></td>
		</tr>
	</tbody>
</table>

fat-water phantoms for magnetic resonance imaging validation: a flexible and scalable protocol

As new techniques are developed to image adipose tissue, methods to validate such protocols are becoming increasingly important. Phantoms, experimental replicas of a tissue or organ of interest, provide a low cost, flexible solution. However, without access to expensive and specialized equipment, constructing stable phantoms with high fat fractions (e.g., &#62;50% fat fraction levels such as those seen in brown adipose tissue) can be difficult due to the hydrophobic nature of lipids. This work presents a detailed, low cost protocol for creating 5x 100 mL&#160;phantoms with fat fractions of 0%, 25%, 50%, 75%, and 100% using basic lab supplies (hotplate, beakers, etc.) and easily accessible components (distilled water, agar, water-soluble surfactant, sodium benzoate, gadolinium-diethylenetriaminepentacetate (DTPA) contrast agent, peanut oil, and oil-soluble surfactant). The protocol was designed to be flexible; it can be used to create phantoms with different fat fractions and a wide range of volumes. Phantoms created with this technique were evaluated in the feasibility study that compared the fat fraction values from fat-water magnetic resonance imaging to the target values in the constructed phantoms. This study yielded a concordance correlation coefficient of 0.998 (95% confidence interval: 0.972-1.00). In summary, these studies demonstrate the utility of&#160;fat phantoms for validating adipose tissue imaging techniques across a range of clinically relevant tissues and organs.

If the water solution has been prepared correctly, a small amount of the solution should congeal quickly in a test vial (Figure 1, left). If the solution separates (Figure 1, right), the solution should be prepared again (as instructed in step 3.8 of the protocol). If the emulsion separates (examples in Figure 2, left and right), the phantom is not v...

Watch this Scientific Journal Video about Fat-Water Phantoms for Magnetic Resonance Imaging Validation: A Flexible and Scalable Protocol at JoVE.com

Fat-Water Phantoms for Magnetic Resonance Imaging Validation: A Flexible and Scalable Protocol

The purpose of this work is to describe a protocol for creating a practical fat-water phantom that can be customized to produce phantoms with varying fat percentages and volumes.

As new techniques are developed to image adipose tissue, methods to validate such protocols are becoming increasingly important. Phantoms, experimental ...

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