Name: Agarose-based Tissue Mimicking Optical Phantoms for Diffuse Reflectance Spectroscopy (Video) | JoVE Journal
Uploaded: 2018-08-22T14:00:00.000+00:00
Duration: 9 min 25 s
Description: Learn to create agarose-based tissue-mimicking optical phantoms for diffuse reflectance spectroscopy in Biomedical Optics. This method enables accurate characterization of optical properties using integrating sphere systems.

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5 minutes

Part of this work was supported by a Grant-in-Aid for Scientific Research (C) from the Japanese Society for the Promotion of Science (25350520, 22500401, 15K06105) and the US-ARMY ITC-PAC Research and Development Project (FA5209-15-P-0175, FA5209-16-P-0132).

1. Construction of a Conventional Diffuse Reflectance and Total Transmittance Spectroscopic System
Note: Construct the measuring systems for the diffuse reflectance and total transmittance spectra using a broadband white light source, a light guide, an achromatic lens, an integrating sphere, a sample holder, an optical fiber, and a multi-channel spectrometer. The role of the light trap is to remove the specular reflection component from the reflectance spectrum. The sample holder of the integrating sphere consists of a mounting plate and a dovetail and spring-loaded clamp assembly that holds the sample against the port. The dovetail and spring-loaded clamp assembly are removed from the sample holder and a hand-made cubic pedestal of polystyrene foam is attached to the mounting plate instead. The layouts of the optical components, shown in Figure 1a and 1b, can be referred to for the construction procedure for the diffuse reflectance measurements and the total transmittance measurements, respectively.
<ol>
	<li>Connect the spectrometer and a personal computer using the universal serial bus (USB) cable provided.</li>
	<li>Attach the port adapter to a detector port of the integrating sphere. Connect the spectrometer and the port adapter of the integrating sphere using an optical fiber. Connect the 150 W halogen lamp light source and the light guide.</li>
	<li>Attach the sample holder to a sample port of the integrating sphere. Attach the light trap to an appropriate port of the integrating sphere when performing the diffuse reflectance measurements. Turn on the halogen lamp light source to illuminate a sample via the light guide and the achromatic lens.</li>
	<li>Open the operating software of the spectrometer.</li>
</ol>
2. Preparation of an Acrylic Mold
Note: An acrylic mold that consists of two rectangular acrylic pieces and a U-shaped acrylic piece is constructed to create a monolayer gel phantom. Figure 2 can be referred to for this construction procedure.
<ol>
	<li>Cut out the two rectangular acrylic pieces from a 2-mm-thick acrylic plate to an optional size.</li>
	<li>Cut out an acrylic piece from a 1-mm-thick acrylic plate to an optional size. Cut the 1-mm-thick acrylic piece so that it becomes a U-shaped piece to be used for the mold to make 1-mm thick epidermal phantoms.</li>
	<li>Cut out an acrylic piece from a 5-mm-thick acrylic plate to an optional size. Cut the 5-mm-thick acrylic piece so that it becomes a U-shaped piece to be used as a mold to make 5-mm thick dermal phantoms.</li>
	<li>Remove any burrs from each acrylic piece using a metal file.</li>
	<li>Make the epidermal phantom mold by holding the 1-mm-thick U-shaped piece with the two 2-mm-thick acrylic pieces and fixing them with five foldback clips.</li>
	<li>Make the dermal phantom mold by holding the 5-mm-thick U-shaped piece with the two 2-mm-thick acrylic pieces and fixing them with five foldback clips.</li>
</ol>
3. Preparation of Base Material
<ol>
	<li>Put 500 mL of standard saline with 0.9% (w/v) NaCl in a pod. Slowly add 5 g of agarose powder while stirring the mixture to avoid clumping.</li>
	<li>Heat the mixture of agarose powder and saline by an electric cooking heater with a 1,000 W power setting for 5 min.</li>
	<li>Once the mixture boils, keep the mixture on low heat for 3 min.</li>
	<li>Cool the mixture to a temperature of about 70 &#176;C. Then pour the mixture into a container and keep it in a constant temperature bath at 60 &#176;C for 30 min before making a phantom.</li>
</ol>
4. Preparation of Skin-mimicking Optical Phantoms
Note: A coffee solution is used to mimic the absorption spectrum of melanin. The coffee solution contains a brown pigment called melanoidin. The absorption spectrum of melanoidin has been reported to be similar to that of melanin10.
<ol>
	<li>Prepare an epidermal phantom
	<ol>
		<li>Pour 100 mL of pure water into the coffee maker reservoir. Place a filter in the coffee maker basket. Add 24 g of ground coffee into the filter. Turn on the coffee maker and press the brew button to begin brewing.</li>
		<li>Put 4 mL of brewed coffee and 16 mL of saline in a glass bottle to make a coffee solution.</li>
		<li>Put 5 mL of lipid emulsion (e.g., intralipid 10%) and 10 mL of the coffee solution into a transparent plastic cup. Slowly add 35 mL of the base material to this mixture while stirring.</li>
		<li>Aspirate the mixture into a syringe and inject it slowly into the epidermal phantom mold while avoiding any bubble formation. Cool the acrylic mold containing the mixture at 5 &#176;C for 20 min.</li>
		<li>Remove the foldback clips from the mold. Slide one of the acrylic pieces outward and remove it from the mold. Take the 1-mm-thick solidified gel phantom out of the mold and cut it to the desired size using a surgical scalpel.</li>
		<li>Place and hold the gel phantom between two slide glasses.</li>
	</ol>
	</li>
	<li>Prepare a dermal phantom containing oxygenated blood
	<ol>
		<li>Take 5.0 mL of lipid emulsion and 0.4 mL of whole equine blood with 45%-hematocrit and put into a transparent plastic cup. Slowly add 44.6 mL of the base material while stirring the mixture.</li>
		<li>Aspirate the mixture into a syringe and inject it slowly into the dermal phantom mold while avoiding any bubble formation. Cool the acrylic mold containing the mixture at 5 &#176;C for 20 min.</li>
		<li>Remove the foldback clips from the mold. Slide one of the acrylic pieces outward and remove it from the mold. Take the 5-mm-thick solidified gel phantom out of the mold and cut it to the desired size using a surgical scalpel.</li>
		<li>Place and hold the gel phantom between two slide glasses.</li>
	</ol>
	</li>
	<li>Prepare a dermal phantom containing deoxygenated blood
	<ol>
		<li>Put a dermal gel phantom containing oxygenated blood (from step 4.2.3) on a glass dish.</li>
		<li>Dissolve 1 g of sodium dithionite (Na2S2O4) into 20 mL of saline in a glass bottle.</li>
		<li>Add 0.05 g/mL of Na2S2O4 solution onto the phantom using a syringe to deoxygenate the blood in the phantom.</li>
		<li>Place and hold the phantom between two slide glasses to prevent it drying out.</li>
	</ol>
	</li>
	<li>Prepare a two-layered phantom
	<ol>
		<li>Drop 0.1 mL of saline onto a dermal phantom to ensure optical coupling between the epidermal and dermal layers. Place the epidermal phantom on the dermal phantom.</li>
		<li>If any air bubbles are present between the layers, push them out by stroking the surface of the two-layered phantom with a fingertip.</li>
		<li>Hold the two-layered phantom between two slide glasses to prevent it drying out.</li>
	</ol>
	</li>
</ol>
5. Acquisition of the Diffuse Reflectance Spectra
<ol>
	<li>Acquisition of dark spectra 
	Note: The charge-coupled device (CCD) sensor in the spectrometer can estimate light intensity based on an electrical signal generated in response to incident light. However, there is dark noise37 which is independent of the signals generated by photons but is dependent on the device temperature, even if the sensor does not detect the light. To accurately measure the spectral intensity of light, the dark current signal should be measured as a dark spectrum and then subtracted from the sample spectrum. The dark spectrum is a spectrum taken with the light path blocked.
	<ol>
		<li>Position the integrating sphere at an optimal position for the diffuse reflectance measurements (Figure 1a).</li>
		<li>Turn off the halogen lamp light source. Block the light path to the spectrometer using a port plug or a shielding plate.</li>
		<li>Select the Store dark command from the file menu to store a dark spectrum.</li>
		<li>Select the Subtract dark spectrum command from the file menu to subtract the dark spectrum from the measured sample spectrum (see below).</li>
	</ol>
	</li>
	<li>Acquisition of reference spectra 
	Note: The optical properties of the components used in this experiment, such as the light source, light guide, achromatic lens, optical fiber, and spectrometer, have their own wavelength-dependences. Therefore, the spectral intensity of light passed through these optical components should be measured as a reference spectrum. For the measurement of a diffuse reflectance spectrum, the reference spectrum is a spectrum taken with a standard white diffuser illuminated with the light from the light source.
	<ol>
		<li>Turn on the halogen lamp light source by pressing the power button. Warm up the lamp for at least 10 min before acquiring a reference spectrum.</li>
		<li>Place a standard white diffuser (e.g., Spectralon) at the sample port of the integrating sphere.</li>
		<li>Adjust the integration time of the spectrometer by selecting the suitable value from the drop-down list in the spectrometer operating software so that the peak signal intensity is approximately 75% of the spectrometer intensity maximum.</li>
		<li>Select the Store reference command from the file menu to store a reference spectrum.</li>
	</ol>
	</li>
	<li>Acquisition of sample spectra 
	Note: A spectrum of the diffuse reflectance of the sample is acquired and saved on the hard drive of a personal computer using the same acquisition conditions.
	<ol>
		<li>Place the epidermal phantom sandwiched by the two slide glasses at the sample port. Select the Save command from the file menu to save a diffuse reflectance spectrum to a file.</li>
		<li>Repeat step 5.3.1 for the dermal and two-layered phantoms.</li>
	</ol>
	</li>
</ol>
6. Acquisition of the Total Transmittance Spectrum
<ol>
	<li>Acquisition of dark spectra 
	Note: The sensor in the spectrometer can estimate light intensity based on an electrical signal generated in response to incident light. However, there is dark noise which is independent of the signals generated by photons but is dependent on the device temperature, even if the sensor does not detect the light. To accurately measure the spectral intensity of light, the dark current signal should be measured as a dark spectrum and then subtracted from the sample spectrum. The dark spectrum is a spectrum taken with the light path blocked.
	<ol>
		<li>Position the integrating sphere at an optimal position for the total transmittance measurements (Figure 1b).</li>
		<li>Remove the light trap from the port of the integrating sphere and attach a port plug to the port.</li>
		<li>Turn off the halogen lamp light source. Block the light path to the integrating sphere using a port plug or shielding plate.</li>
		<li>Select the Store dark command from the file menu to store a dark spectrum.</li>
		<li>Select the Subtract dark spectrum command from the file menu to subtract the dark spectrum from the measured sample spectrum (see below).</li>
	</ol>
	</li>
	<li>Acquisition of reference spectra 
	Note: The optical properties of the components used in this experiment, such as the light source, light guide, achromatic lens, optical fiber, and spectrometer, have their own wavelength-dependences. Therefore, the spectral intensity of the light passed through these components should be measured as a reference spectrum. For the measurement of the total transmittance spectrum, the reference spectrum is a spectrum taken when the light from the light source is directly entering the integrating sphere through the sample port.
	<ol>
		<li>Turn on the halogen lamp light source by pressing the power button. Warm up the lamp for at least 10 min before acquiring a reference spectrum.</li>
		<li>Regulate the integration time of the spectrometer by selecting the suitable value from the drop-down list of integration times in the operating software of the spectrometer so that the greatest light intensity shows a signal that is approximately 75% of the maximum values.</li>
		<li>Select the Store reference command from the file menu to store a reference spectrum.</li>
	</ol>
	</li>
	<li>Acquisition of sample spectra 
	Note: The spectrum of the total transmittance of the sample is acquired and saved on the hard drive of a personal computer using the same acquisition conditions.
	<ol>
		<li>Place the epidermal phantom sandwiched by the two slide glasses at the sample port. Select the Save command from the file menu to save a total transmittance spectrum to a file.</li>
		<li>Repeat step 6.3.1 for the dermal and two-layered phantoms.</li>
	</ol>
	</li>
</ol>
7. Estimating the Absorption and Light-scattering Properties
Note: A set of the diffuse reflectance spectrum and the total transmittance spectrum is saved to the hard drive of a personal computer and analyzed offline. An inverse Monte Carlo simulation8,38,39,40 is then performed to estimate the absorption coefficient spectrum &#181;a(&#955;) and the reduced scattering coefficient spectrum &#181;s&#8217;(&#955;). In this inverse Monte Carlo simulation, the estimated scattering coefficient &#181;s, under the assumption that the anisotropy factor g is 0, is regarded as the reduced scattering coefficient &#181;s&#8217;. Both the reflectance and the transmittance data are used for a single simulation run. The detailed algorithm used in this protocol has been reported in previous literature8,39. We estimated the absorption coefficient spectrum &#181;a(&#955;) and the reduced scattering coefficient spectrum &#181;s&#8217;(&#955;) of an epidermal layer from a set of the diffuse reflectance spectrum and the total transmittance spectrum obtained from the epidermal layer. In the same way, we estimated &#181;a(&#955;) and &#181;s&#8217;(&#955;) of a dermal layer from a set of the diffuse reflectance spectrum and the total transmittance spectrum obtained from the dermal layer.
<ol>
	<li>Open an input file for the Monte Carlo simulation.</li>
	<li>Fill in the values of the measured diffuse reflectance and the total transmittance at the specific wavelength range from 400 to 700 nm at 10 nm-intervals in the input data file. Fill in the value of the phantom thickness in the input data file.</li>
	<li>Set the refractive index n of a layer to be an appropriate value in the input data file (e.g., n = 1.33 at 550 nm). Set the value of the anisotropy factor g to be 0 in the input data file.</li>
	<li>Set the initial values of the absorption coefficient &#181;a and the scattering coefficient &#181;s to be the appropriate values in the input data file (e.g., &#181;a = 0.01, &#181;s = 0.1).</li>
	<li>Execute the inverse Monte Carlo simulation program.</li>
	<li>Type the input file name and then run the simulation.</li>
	<li>Open the output file and check the final values of &#181;a and &#181;s after the iterative simulation is terminated.</li>
	<li>Repeat steps 7.1 - 7.7 for other desired wavelengths.</li>
</ol>

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<li>Ebert, B., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Near-infrared+fluorescent+dyes+for+enhanced+contrast+in+optical+mammography%3A+phantom+experiments%5BTitle%5D)+AND+%22Journal+of+Biomedical+Optics%22%5BJournal%5D)">Near-infrared fluorescent dyes for enhanced contrast in optical mammography: phantom experiments.</a> Journal of Biomedical Optics. 6 (2), 134-140 (2001).</li>
<li>Sukowski, U., Schubert, F., Grosenick, D., Rinneberg, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Preparation+of+solid+phantoms+with+defined+scattering+and+absorption+properties+for+optical+tomography%5BTitle%5D)+AND+%22Physics+in+Medicine+%26+Biology%22%5BJournal%5D)">Preparation of solid phantoms with defined scattering and absorption properties for optical tomography.</a> Physics in Medicine & Biology. 41, 1823-1844 (1996).</li>
<li>Yoshida, K., Nishidate, I., Ishizuka, T., Kawauchi, S., Sato, S., Sato, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Multispectral+imaging+of+absorption+and+scattering+properties+of+in+vivo+exposed+rat+brain+using+a+digital+red-green-blue+camera%5BTitle%5D)+AND+%22Journal+of+Biomedical+Optics%22%5BJournal%5D)">Multispectral imaging of absorption and scattering properties of in vivo exposed rat brain using a digital red-green-blue camera.</a> Journal of Biomedical Optics. 20 (5), 051026(2015).</li>
<li>Lo, J. Y., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((A+strategy+for+quantitative+spectral+imaging+of+tissue+absorption+and+scattering+using+light+emitting+diodes+and+photodiodes%5BTitle%5D)+AND+%22Optics+Express%22%5BJournal%5D)">A strategy for quantitative spectral imaging of tissue absorption and scattering using light emitting diodes and photodiodes.</a> Optics Express. 17 (3), 1372-1384 (2009).</li>
<li>Bednov, A., Ulyanov, S., Cheung, C., Yodh, A. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Correlation+properties+of+multiple+scattered+light%3A+implication+to+coherent+diagnostics+of+burned+skin%5BTitle%5D)+AND+%22Journal+of+Biomedical+Optics%22%5BJournal%5D)">Correlation properties of multiple scattered light: implication to coherent diagnostics of burned skin.</a> Journal of Biomedical Optics. 9 (2), 347-352 (2004).</li>
<li>Hull, E. L., Nichols, M. G., Foster, T. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Quantitative+broadband+near-infrared+spectroscopy+of+tissue-simulating+phantoms+containing+erythrocytes%5BTitle%5D)+AND+%22Physics+in+Medicine+%26+Biology%22%5BJournal%5D)">Quantitative broadband near-infrared spectroscopy of tissue-simulating phantoms containing erythrocytes.</a> Physics in Medicine & Biology. 43 (11), 3381-3404 (1998).</li>
<li>Kienle, A., Patterson, M. S., Ott, L., Steiner, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Determination+of+the+scattering+coefficient+and+the+anisotropy+factor+from+laser+Doppler+spectra+of+liquids+including+blood%5BTitle%5D)+AND+%22Applied+Optics%22%5BJournal%5D)">Determination of the scattering coefficient and the anisotropy factor from laser Doppler spectra of liquids including blood.</a> Applied Optics. 35 (19), 3404-3412 (1996).</li>
<li>Srinivasan, S., Pogue, B. W., Jiang, S., Dehghani, H., Paulsen, K. D. <a target="_blank" href="https://www.osapublishing.org/ao/abstract.cfm?uri=ao-44-10-1858">Spectrally constrained chromophore and scattering NIR tomography improves quantification and robustness of reconstruction.</a> Applied Optics. 44 (10), 1858-1869 (2004).</li>
<li>Ocean Optics Inc. Glossary. Dark Noise. , Available from:
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<li>Friebel, M., Roggan, A., Müller, G., Meinke, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Determination+of+optical+properties+of+human+blood+in+the+spectral+range+250+to+1100+nm+using+Monte+Carlo+simulations+with+hematocrit-dependent+effective+scattering+phase+functions%5BTitle%5D)+AND+%22Journal+of+Biomedical+Optics%22%5BJournal%5D)">Determination of optical properties of human blood in the spectral range 250 to 1100 nm using Monte Carlo simulations with hematocrit-dependent effective scattering phase functions.</a> Journal of Biomedical Optics. 11 (3), 34021(2006).</li>
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<li>Friebel, M., Helfmann, J., Netz, U., Meinke, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Influence+of+oxygen+saturation+on+the+optical+scattering+properties+of+human+red+blood+cells+in+the+spectral+range+250+to+2000+nm%5BTitle%5D)+AND+%22Journal+of+Biomedical+Optics%22%5BJournal%5D)">Influence of oxygen saturation on the optical scattering properties of human red blood cells in the spectral range 250 to 2000 nm.</a> Journal of Biomedical Optics. 14 (3), 034001(2009).</li>
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<li>Park, J., Ha, M., Yu, M., Jung, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Fabrication+of+various+optical+tissue+phantoms+by+the+spin-coating+method%5BTitle%5D)+AND+%22Journal+of+Biomedical+Optics%22%5BJournal%5D)">Fabrication of various optical tissue phantoms by the spin-coating method.</a> Journal of Biomedical Optics. 21 (6), 065008(2016).</li>
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</ol>

The authors have nothing to disclose.

The most critical step in this protocol is the temperature control of the base material. The temperature to maintain the base material ranged from 58 to 60 &#176;C. If the temperature is more than 70 &#176;C, a denaturation of both the lipid emulsion and the whole blood will occur. As a consequence, the optical properties of the phantom will deteriorate. If the temperature is less than 40 &#176;C, the base material will be ununiformly gelled and, thus, the light scattering and absorption agents will be heterogeneously di...

Optical phantoms are objects mimicking the optical properties of biological tissues and have been widely used in the biomedical optics field. They are designed so that the optical properties, such as light scattering and absorption coefficients, match with those of living human and animal tissues. Optical phantoms are generally used for the following purposes: simulating the light transport in biological tissues, calibrating a newly developed optical system design, evaluating the quality and performance of existing systems, comparing the performance between systems, and validating the ability of the optical methods to quantify the optical properties1,2,3,4,5. Therefore, easy-to-get substances, a simple fabrication process, a high reproducibility, and an optical stability are required for making optical phantoms.
Various types of optical phantoms with different base materials such as aqueous suspension6, gelatin gel7, agarose gel8,9,10, polyacrylamide gel11, resin12,13,14,15,16, and room-temperature-vulcanizing silicone17 have been reported in previous literature. It has been reported that gelatin- and alginate-based gels are useful for optical phantoms with heterogeneous structures18. Alginate phantoms have a suitable mechanical and thermal stability for evaluating photothermal effects such as laser ablation studies and laser-based hyperthermia studies18. Agarose gels have the ability to fabricate heterogeneous structures, and their mechanical and physical properties are stable for a long time18. High-purity agarose gels have a very low turbidity and a weak optical absorption. Therefore, optical properties of agarose-based phantoms could easily be designed with the appropriate light scattering and absorbing agents. Recently, styrene-ethylene-butylene-styrene (SEBS) block copolymers19 and polyvinyl chloride (PVC) gels20 have been reported as interesting phantom materials for optical and photoacoustic techniques.
Polymer microspheres7,12,21,22, titanium oxide powder1, and lipid emulsions23,24,25,26 such as milk and lipid emulsion are used as light scattering agents, whereas black ink27,28 and molecular dyes29,30 are used as light absorbers. Diffuse reflectance spectra of most living organs are dominated by the absorption of oxygenated and deoxygenated hemoglobin in red blood cells. Therefore, hemoglobin solutions31,32 and whole blood8,9,10,33,36 are often used as light absorbers in the phantoms for a diffuse reflectance spectroscopy and multispectral imaging.
The method described in this article is used to create an optical phantom mimicking the light transport in biological tissues and to characterize its optical properties. As an example, a two-layered optical phantom mimicking optical properties of human skin tissue is demonstrated. The advantages of this method over alternative techniques are the ability to represent diffuse reflectance spectra of living biological tissues in the visible to near-infrared wavelength region, as well as the simplicity to make it, using easily available materials and conventional optical instruments. Therefore, the optical phantoms made by this method will be useful for the development of optical methods based on diffuse reflectance spectroscopy and multispectral imaging.

<table><tbody><tr><td>150-W halogen-lamp light source</td><td>Hayashi Watch Works Co., Ltd, Tokyo, Japan</td><td>LA-150SAE</td><td></td></tr><tr><td>Light guide</td><td>Hayashi Watch Works Co., Ltd, Tokyo, Japan</td><td>LGC1-5L1000</td><td></td></tr><tr><td>Integrating Sphere</td><td>Labsphere Incorporated, North Sutton, NH, USA</td><td>RT-060-SF</td><td></td></tr><tr><td>Port adapter</td><td>Labsphere Incorporated, North Sutton, NH, USA</td><td>PA-050-SMA-SF</td><td></td></tr><tr><td>Light trap</td><td>Labsphere Incorporated, North Sutton, NH, USA</td><td>LTRP-100-C</td><td></td></tr><tr><td>Spectralon white standard with 99% diffuse reflectance</td><td>Labsphere Incorporated, North Sutton, NH, USA</td><td>SRS-99-020</td><td></td></tr><tr><td>Optical fiber</td><td>Ocean Optics Inc., Dunedin, Florida, USA</td><td>P400-2-VIS-NIR</td><td></td></tr><tr><td>Miniature Fiber Optic Spectrometer</td><td>Ocean Optics Inc., Dunedin, Florida, USA</td><td>USB2000</td><td></td></tr><tr><td>Achromatic lens</td><td>Chuo Precision Industrial Co.,Ltd, Tokyo, Japan</td><td>ACL-50-75M</td><td></td></tr><tr><td>Intralipid</td><td>Fresenius Kabi AB, Uppsala, Sweden</td><td>Intralipid 10%</td><td></td></tr><tr><td>Coffee&lt;br/&gt; (Blendy Mocha Blend Regular Coffee)</td><td>Ajinomoto AGF, Inc. Tokyo, Japan</td><td>Unavailable</td><td></td></tr><tr><td>Whole blood</td><td>Nippon Bio-Test Laboratories Inc. Saitama, Japan</td><td>0103-2</td><td></td></tr><tr><td>Agarose</td><td>Nippon Genetics Co., Ltd, Tokyo, Japan</td><td>NE-AG02</td><td></td></tr><tr><td>Cooking heater</td><td>TOSHIBA CORPORATION Tokyo, Japan</td><td>HP-103K</td><td></td></tr></tbody></table>

agarose-based tissue mimicking optical phantoms for diffuse reflectance spectroscopy

Agarose-based Tissue Mimicking Optical Phantoms for Diffuse Reflectance Spectroscopy

This protocol describes how to make agarose-based tissue-mimicking phantoms and demonstrates how to determine their optical properties using a conventional optical system with an integrating sphere. Measuring systems for the acquisition of the diffuse reflectance and total transmittance spectra are constructed with a broadband white light source, a light guide, an achromatic lens, an integrating sphere, a sample holder, an optical fiber probe, and a multi-channel spectrometer. An acrylic mold consisting of two rectangular acrylic pieces and a U-shaped acrylic piece is constructed to create an epidermal phantom and a dermal phantom with whole blood. The application of a sodium dithionite (Na2S2O4) solution to the dermal phantom enables the researcher to deoxygenate hemoglobin in red blood cells distributed in the dermal phantom. The inverse Monte Carlo simulation with the diffuse reflectance and total transmittance spectra measured by a spectrometer with an integrating sphere is performed to determine the absorption coefficient spectrum &#181;a(&#955;) and the reduced scattering coefficient spectrum &#181;s&#39;(&#955;) of each layer phantom. A two-layered phantom mimicking the diffuse reflectance of human skin tissue is also demonstrated by piling up the epidermal phantom on the dermal phantom.

Figure 3 shows the representative estimated spectra of the reduced scattering coefficient and the absorption coefficient for the epidermal phantom and dermal phantom. The results shown in Figure 3 are the averages of ten measurements of both reflectance and transmittance spectra. The reduced scattering coefficient µs' has a broad scattering spectrum, exhibiting a higher magnitude at shorter wavelength...

Here, we demonstrate how agarose-based tissue-mimicking optical phantoms are made and how their optical properties are determined using a conventional optical system with an integrating sphere.

This protocol describes how to make agarose-based tissue-mimicking phantoms and demonstrates how to determine their optical properties using a ...

agarose-based-tissue-mimicking-optical-phantoms-for-diffuse

Research

JoVE Journal

Bioengineering

Near-infrared fluorescence (NIRF) imaging may improve therapeutic outcome of breast cancer surgery by enabling intraoperative tumor localization and evaluation of surgical margin status. Using tissue-simulating breast phantoms containing fluorescent tumor-simulating inclusions, potential clinical applications of NIRF imaging in breast cancer patients can be assessed for standardization and training purposes.

Tissue-simulating Phantoms for Assessing Potential Near-infrared Fluorescence Imaging Applications in Breast Cancer Surgery

The assembly and use of a multimodal microendoscope is described which can co-register superficial tissue image data with tissue physiological parameters including hemoglobin concentration, melanin concentration, and oxygen saturation. This technique can be useful for evaluating tissue structure and perfusion, and can be optimized for individual needs of the investigator.

Multimodal Imaging and Spectroscopy Fiber-bundle Microendoscopy Platform for Non-invasive, In Vivo Tissue Analysis

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

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

This protocol describes how the use of diffuse polarization spectroscopy can improve the clinical usefulness of the capillary refill test. We suggest a more detailed analysis of the course of the capillary refill in healthy volunteers using diffuse reflectance spectroscopy videos and new informatic endpoints.

Diffuse Reflectance Spectroscopy: Getting the Capillary Refill Test Under One's Thumb

Optical tissue phantoms are essential tools for calibration and characterization of optical imaging systems and validation of theoretical models. This article details a method for phantom fabrication that includes replication of tissue optical properties and three-dimensional tissue structure.

Fabrication and Characterization of Optical Tissue Phantoms Containing Macrostructure

Spin coating, polyjet printing, and fused deposition modeling are integrated to produce multilayered heterogeneous phantoms that simulate structural and functional properties of biological tissue.

Multimodal 3D Printing of Phantoms to Simulate Biological Tissue

This protocol describes the fabrication of a stable, biologically relevant phantom material for optical and acoustic biomedical imaging applications, featuring independently tunable acoustic and optical properties.

A Stable Phantom Material for Optical and Acoustic Imaging

This article presents the protocol for preparing tumor-tissue phantoms that replicate optical properties for plasmonic photothermal therapy. It details phantom preparation, photothermal evaluations, and validation of the developed numerical model based on photothermal temperature measurements for assessing therapeutic parameters, offering an ethical, cost-effective alternative to in vivo studies for preliminary testing.

Plasmonic Photothermal Cancer Therapy: Nanoparticle-embedded Tumor-tissue-mimicking Phantoms for Visualizing Photothermal Temperature Distribution

Quantitative, high-throughput, real-time, and label-free biomolecular detection (DNA, protein, etc.) on SiO2 surfaces can be achieved using a simple interferometric technique which relies on LED illumination, minimal optical components, and a camera.  The Interferometric Reflectance Imaging Sensor (IRIS) is inexpensive, simple to use, and amenable to microarray formats.

Biomolecular Detection employing the Interferometric Reflectance Imaging Sensor (IRIS)

Raman spectroscopy is a suitable technique for the non-contact, label-free analysis of living cells, tissue-engineered constructs and native tissues. Source-specific spectral fingerprints can be generated and analyzed using multivariate analysis.

Non-contact, Label-free Monitoring of Cells and Extracellular Matrix using Raman Spectroscopy

Learn to create agarose-based tissue-mimicking optical phantoms for diffuse reflectance spectroscopy in Biomedical Optics. This method enables accurate characterization of optical properties using integrating sphere systems.

Agarose-based Tissue Mimicking Optical Phantoms for Diffuse Reflectance Spectroscopy

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