The research leading to these results has received funding from the European Research Council under grant agreement ERC-2010-StG-260991.

La procédure détaillée pour fonctionner avec la sonde acousto-optique à main volumétrique est décrit ci-dessous. Cette procédure est effectuée selon la réglementation en ce qui concerne l&#39;établissement approuvés expériences animales et humaines. 1. Préparation du système <ol><li> Mettre en marche le laser pendant une période d&#39;échauffement de ~ 15 min avant l&#39;opération de stabilisation du faisceau de lumière de sortie. </li><li> Placer la partie de l&#39;eau renfermant de la membrane d&#39;isolement qui est en contact avec la peau (Figure 1). REMARQUE: La distance entre la membrane d&#39;isolement (en contact avec la peau) et la région de sensibilité maximale du capteur (centre du palpeur sphérique) définit la profondeur d&#39;imagerie efficace. </li><li> Compléter le volume total d&#39;environ 100 ml entre la membrane isolante et la surface du transducteur avec de l&#39;eau désionisée à l&#39;aide d&#39;une pompe. </li><li> Assurez-vous qu&#39;aucun des bulles de fuite d&#39;eau ni air sont present. Sinon, éviter les bulles d&#39;air en fournissant recirculation de l&#39;eau. </li><li> Réaliser les essais à la température ambiante et assurer le milieu de couplage (eau) est maintenue à cette température. </li></ol> 2. Préparation d&#39;imagerie <ol><li> Préparation d&#39;imagerie humain. </li><li> Enlever les poils de la partie à être imagées avec une lotion d&#39;épilation afin d&#39;éviter un fond indésirable dans les images (cette étape est facultative). </li><li> Appliquer le gel à ultrasons à la peau autour de la région à imager afin de fournir un couplage acoustique efficace. Placer la sonde optoacoustique dans la région d&#39;intérêt. Veiller à ce que les bulles d&#39;air sont présents dans le gel de couplage ultrasonore. </li><li> préparation d&#39;imagerie animale. </li><li> Veiller à ce que les procédures de soins et expérimentales sur les animaux sont en accord avec les règles et les règlements institutionnels et gouvernementaux. </li><li> Retirer la fourrure de l&#39;animal dans la zone à imager avec une lotion de rasage. Protéger les yeux de l&#39;animal avecvétérinaire pommade, qui prévient le dessèchement et les dommages causés par l&#39;exposition à un rayonnement laser intense pulsée. </li><li> Anesthésier l&#39;animal à l&#39;aide d&#39;une injection intrapéritonéale (IP) de kétamine / xylazine (100 mg / kg kg de kétamine + 5 mg / kg KG xylazine) avant l&#39;expérience ou une anesthésie isoflurane (2-3% (en volume) avec 0,9 l / min débit de gaz) pendant l&#39;expérience. Confirmer anesthésie en vérifiant le réflexe de la patte postérieure de l&#39;animal. </li><li> Appliquer le gel à ultrasons à la peau autour de la région à imager afin de fournir un couplage acoustique efficace et placer la sonde optoacoustique dans la région d&#39;intérêt. Veiller à ce que les bulles d&#39;air sont présents dans le gel de couplage ultrasonore. </li></ol> 3. Pré-affichage en mode de fonctionnement <ol><li> Mettre en place la longueur d&#39;onde (s) de formation d&#39;image entre 690 nm et 900 nm et le taux de répétition des impulsions entre 10 et 50 Hz. Sélectionner les paramètres du système d&#39;acquisition de données acoustiques - 1 MQimpédance d&#39;entrée. Acquérir 2030 échantillons pour chaque impulsion de laser à un taux de 40 Mégaéchantillons par seconde et 12 bits résolution verticale échantillonnage. Déclencher l&#39;acquisition de la sortie Q-switch du laser. </li><li> Veiller à ce que l&#39;opérateur et le patient utilisent des lunettes de protection adaptées à la longueur d&#39;onde (s) d&#39;excitation optique. Régler la puissance du laser de sorte que l&#39;influence de la lumière à la surface de tissu est maintenue en dessous de 20 mJ / cm 2 lors de l&#39;expérience pour des longueurs d&#39;onde du proche infrarouge de manière à satisfaire des limites d&#39;exposition de sécurité pour les expériences de l&#39;homme 19 et pour éviter la fatigue thermique et de lésions de la peau chez les animaux . </li><li> Démarrer le logiciel de pré-vision avec une mise en oeuvre de GPU d&#39;algorithmes de traitement pour permettre la visualisation d&#39;images tridimensionnelles à une vitesse de défilement correspondant à la vitesse de répétition des impulsions du laser. </li><li> Déplacer la sonde et / ou l&#39;objet à imager afin d&#39;optimiser les performances de visualisation et de localiser les structures d&#39;intérêt. </li></ol> 4. Acquisition de données <ol><li> L&#39;acquisition des données pour le mode de balayage (à main). </li><li> Si besoin est, d&#39;injecter un produit de contraste avant l&#39;acquisition d&#39;enrichir le contraste dans la région d&#39;intérêt. NOTE: Dans nos expériences nous ont pas effectué imagerie humaine contraste amélioré. Cependant, divers agents de contraste peuvent être potentiellement utilisés dans ce but. Vert d&#39;indocyanine (ICG), est un exemple d&#39;agent de contraste optique cliniquement approuvé qui peut être utilisé pour l&#39;amélioration du contraste à dose maximale recommandée de 2 mg / kg de poids corporel chez l&#39;adulte. </li><li> Lancer le matériel d&#39;acquisition de données avec les paramètres décrits dans le maintien de la 3.1 l&#39;exécution du logiciel de prévisualisation. Déplacez doucement la sonde autour de la région imagée pour suivre les structures d&#39;intérêt. REMARQUE: lorsque les images à de multiples longueurs d&#39;onde laser sont acquis simultanément, la vitesse de déplacement de la sonde dans le mode portatif doit être diminué de façon significative (de préférence inférieure à 2 mm / spour un taux de répétition des impulsions laser de 50 Hz) afin d&#39;éviter les artefacts relative au mouvement dans les images spectrale non mélangés. </li><li> L&#39;acquisition des données en mode stationnaire. </li><li> Montez l&#39;objet imagé (ex., Animaux) et la sonde à main sur le support et lancer l&#39;acquisition avec les paramètres décrits à la section 3.1 de maintien de l&#39;exécution du logiciel pré-vue. </li><li> Maintenir la sonde optoacoustique et la partie de formation d&#39;image dans la même position pendant l&#39;expérience pour visualiser événements biologiques dynamiques de la région d&#39;intérêt. </li><li> Injecter un agent de contraste pour suivre sa distribution dynamique dans la région d&#39;intérêt. REMARQUE: Dans nos expériences, des souris, vert d&#39;indocyanine (ICG) a été utilisé pour l&#39;augmentation du contraste. En règle générale, une quantité de 10 nmoles ou 0,4 mg / kg d&#39;ICG doit être introduit dans la circulation sanguine de la souris afin de créer un contraste détectable avec optoacoustics multi-spectrales in vivo. REMARQUE: L&#39;agent de contraste doit êtreapprouvé pour un usage humain et / ou animal par l&#39;autorité respective. </li></ol> 5. Finaliser l&#39;expérience <ol><li> Arrêtez le laser. </li><li> Retirer la sonde acousto-optique de la région imagée. Pour l&#39;étude des animaux, arrêter l&#39;alimentation de l&#39;anesthésie. </li><li> Placez l&#39;animal sous un chauffage à infrarouge pour garder la chaleur et éviter le contact avec d&#39;autres animaux jusqu&#39;à ce que complètement récupéré de l&#39;anesthésie. Ne pas laisser l&#39;animal sans surveillance lors de la récupération de l&#39;anesthésie. </li></ol> 6. Off-line de traitement des données <ol><li> Chargez le fichier (s) contenant les signaux acousto-optique acquise dans l&#39;application de logiciels utilisés pour le traitement des données. </li><li> Utiliser un algorithme de reconstruction pour obtenir une matrice à trois dimensions correspondant à une image volumétrique de l&#39;absorption optique pour chaque trame et chaque longueur d&#39;onde. NOTE: Pour la reconstruction, il est préférable d&#39;utiliser un algorithme tenant compte de facteurs de distorsion, comme HeterogeneLITÉS et l&#39;atténuation de l&#39;objet imagé, les effets de bande passante finale et forme géométrique des éléments de détection et de légères variations de fluence, afin d&#39;obtenir une représentation plus quantitative de la distribution de l&#39;énergie absorbée. </li><li> Utiliser un algorithme de déconvolution pour obtenir, à partir de chaque trame multi-longueur d&#39;onde, une nouvelle série de tableaux de la matrice en trois dimensions représentant l&#39;absorption optique pour chaque substance absorbante présente dans l&#39;échantillon. </li><li> Si nécessaire, traiter en outre les matrices de la matrice représentant la distribution de l&#39;absorption optique pour faciliter la visualisation et la lecture des paramètres biologiques pertinents. </li></ol>

<ol>
	<li>Stritzker, J., 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=Vaccinia+virus-mediated+melanin+production+allows+MR+and+optoacoustic+deep+tissue+imaging+and+laser-induced+thermotherapy+of+cancer.">Vaccinia virus-mediated melanin production allows MR and optoacoustic deep tissue imaging and laser-induced thermotherapy of cancer.</a> Proceedings of the National Academy of Sciences of the United States of America. 110 (9), 3316-3320 (2013).</li><li>Herzog, E., 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=Optical+Imaging+of+Cancer+Heterogeneity+with+Multispectral+Optoacoustic+Tomography.">Optical Imaging of Cancer Heterogeneity with Multispectral Optoacoustic Tomography.</a> Radiology. 263 (2), 461-468 (2012).</li><li>Johnson, S. P., Laufer, J. G., Zhang, E. Z., Beard, P. C., Pedley, R. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Determination+of+Differential+Tumour+Vascular+Pathophysiology+in+Vivo+by+Photoacoustic+Imaging.">Determination of Differential Tumour Vascular Pathophysiology in Vivo by Photoacoustic Imaging.</a> Eur J Cancer. 48, 186-187 (2012).</li><li>Yao, J. J., 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=Noninvasive+photoacoustic+computed+tomography+of+mouse+brain+metabolism+in+vivo.">Noninvasive photoacoustic computed tomography of mouse brain metabolism in vivo.</a> Neuroimage. 64, 257-266 (2013).</li><li>Strohm, E. M., Berndl, E. S. L., Kolios, M. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=High+frequency+label-free+photoacoustic+microscopy+of+single+cells.">High frequency label-free photoacoustic microscopy of single cells.</a> Photoacoustics. 1 (3-4), 49-53 (2013).</li><li>Beard, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Biomedical+photoacoustic+imaging.">Biomedical photoacoustic imaging.</a> Interface Focus. 1, 602-631 (2011).</li><li>Wang, L. H. V., Hu, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Photoacoustic+Tomography:+In+Vivo+Imaging+from+Organelles+to+Organs.">Photoacoustic Tomography: In Vivo Imaging from Organelles to Organs.</a> Science. 335 (6075), 1458-1462 (2012).</li><li>Xiang, L. Z., Wang, B., Ji, L. J., Jiang, H. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=4-D+Photoacoustic+Tomography.">4-D Photoacoustic Tomography.</a> Sci Rep-Uk. 3, 10-1038 (2013).</li><li>Buehler, A., Dean-Ben, X. L., Claussen, J., Ntziachristos, V., Razansky, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Three-dimensional+optoacoustic+tomography+at+video+rate.">Three-dimensional optoacoustic tomography at video rate.</a> Optics express. 20 (20), 22712-22719 (2012).</li><li>Dean-Ben, X. L., Razansky, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Adding+fifth+dimension+to+optoacoustic+imaging:+volumetric+time-resolved+spectrally-enriched+tomography.">Adding fifth dimension to optoacoustic imaging: volumetric time-resolved spectrally-enriched tomography.</a> Light: Science and Applications. 3, (2014).</li><li>Fronheiser, M. 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+optoacoustic+monitoring+and+three-dimensional+mapping+of+a+human+arm+vasculature.">Real-time optoacoustic monitoring and three-dimensional mapping of a human arm vasculature.</a> J Biomed Opt. 15 (2), (2010).</li><li>Buehler, A., Kacprowicz, M., Taruttis, A., Ntziachristos, V. <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+handheld+multispectral+optoacoustic+imaging.">Real-time handheld multispectral optoacoustic imaging.</a> Opt Lett. 38 (9), 1404-1406 (2013).</li><li>Yang, J. M., 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=Simultaneous+functional+photoacoustic+and+ultrasonic+endoscopy+of+internal+organs+in+vivo.">Simultaneous functional photoacoustic and ultrasonic endoscopy of internal organs in vivo.</a> Nat Med. 18 (8), 1297-1302 (2012).</li><li>Kruger, R. A., Lam, R. B., Reinecke, D. R., Del Rio, S. P., Doyle, R. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Photoacoustic+angiography+of+the+breast.">Photoacoustic angiography of the breast.</a> Med Phys. 37 (11), 6096-6100 (2010).</li><li>Heijblom, M., 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=Visualizing+breast+cancer+using+the+Twente+photoacoustic+mammoscope:+What+do+we+learn+from+twelve+new+patient+measurements.">Visualizing breast cancer using the Twente photoacoustic mammoscope: What do we learn from twelve new patient measurements.</a> Optics express. 20 (11), 11582-11597 (2012).</li><li>Dean-Ben, X. L., Razansky, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Portable+spherical+array+probe+for+volumetric+real-time+optoacoustic+imaging+at+centimeter-scale+depths.">Portable spherical array probe for volumetric real-time optoacoustic imaging at centimeter-scale depths.</a> Optics express. 21 (23), 28062-28071 (2013).</li><li>Dean-Ben, X. L., Ozbek, A., Razansky, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Volumetric+real-time+tracking+of+peripheral+human+vasculature+with+GPU-accelerated+three-dimensional+optoacoustic+tomography.">Volumetric real-time tracking of peripheral human vasculature with GPU-accelerated three-dimensional optoacoustic tomography.</a> IEEE transactions on medical imaging. 32 (11), 2050-2055 (2013).</li><li>Dean-Ben, X. L., Razansky, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Functional+optoacoustic+human+angiography+with+handheld+video+rate+three+dimensional+scanner.">Functional optoacoustic human angiography with handheld video rate three dimensional scanner.</a> Photoacoustics. 1 (3-4), 68-73 (2013).</li><li>. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> American National Standards for the Safe Use of Lasers ANSI Z136.1.. , (2000).</li><li>Ntziachristos, V., Razansky, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Molecular+imaging+by+means+of+multispectral+optoacoustic+tomography+(MSOT).">Molecular imaging by means of multispectral optoacoustic tomography (MSOT).</a> Chemical reviews. 110 (5), 2783-2794 (2010).</li><li>Luke, G. P., Yeager, D., Emelianov, S. Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Biomedical+Applications+of+Photoacoustic+Imaging+with+Exogenous+Contrast+Agents.">Biomedical Applications of Photoacoustic Imaging with Exogenous Contrast Agents.</a> Ann Biomed Eng. 40 (2), 422-437 (2012).</li></ol>

The authors have nothing to disclose.

Les avantages uniques offerts par les techniques d&#39;imagerie acousto-optique dans la recherche de petits animaux ont créé une forte motivation pour la traduction de la technologie dans la pratique clinique, avec un certain nombre de diagnostics et les applications de surveillance de traitement par exemple envisagés., Cancer du sein et de la peau, l&#39;inflammation ou les maladies du système vasculaire périphérique. Cependant, par opposition à des souris ou des animaux plus petits, qui peuvent être e...

Acousto-optique (photo-acoustique) imagerie attire de plus en plus l&#39;intérêt des communautés de recherche biologique et médicale, qui se manifeste par le nombre sans cesse croissant de publications couvrant variété de nouvelles applications qui exploitent les avantages uniques offerts par la technologie 1-5. En particulier, la capacité à image distinctive spectrale agents de photo-absorption avec une résolution spatio-temporelle à des profondeurs bien au-delà de la limite de diffusion de la lumière ouvre capacités sans précédent pour l&#39;imagerie fonctionnelle et moléculaire 6-10. En effet, la traduction de la technologie acousto-optique dans la pratique clinique est livré avec des perspectives prometteuses dans le diagnostic et le suivi du traitement de nombreuses maladies. Cependant, la propagation limité de photons dans la dispersion optique et de tissus absorbants et les réponses faibles généralement associés au phénomène optoacoustique limiter la profondeur de la méthode applicable. En conséquence, optoa portatifCoustic sondes ont été tentées pour des parties d&#39;image accessibles depuis l&#39;extérieur du corps de 11,12 alors que les systèmes endoscopiques sont utilisés pour fournir des images de l&#39;intérieur du corps en les insérant par des orifices naturels 13. Certaines parties faible absorption de corps humain, tels que le sein féminin, sont également accessibles par les scanners tomographiques 14,15 acousto-optique. Un intérêt particulier est l&#39;approche à main car il permet un grand polyvalence, de façon similaire à l&#39;échographie. Ici, l&#39;adaptation des ultrasons commune linéaires sondes de tableau pour l&#39;imagerie acousto-optique reste difficile, principalement en raison de différences fondamentales dans les exigences d&#39;imagerie tomographique entre l&#39;échographie et optoacoustics. Bien que les taux de rafraîchissement élevé en échographie norme sont activés par séquentiel émission-réception systèmes utilisant des fréquences de répétition élevé d&#39;impulsions dans la gamme kHz, l&#39;imagerie acousto-optique en temps réel en trois dimensions est obtenue par collecte simultanée de données tomographiques volumétrique d&#39;un seul interrogating impulsion laser. Ainsi, l&#39;imagerie acousto-optique de haute qualité implique l&#39;acquisition de données en trois dimensions de la plus grande angle solide possible autour de l&#39;objet imagé. Récemment, nous avons lancé la première sonde acousto-optique pour portable en trois dimensions (volumétrique) l&#39;imagerie en temps réel 16. Le système est basé sur un réseau bidimensionnel de 256 éléments piézo-électriques disposés sur une surface sphérique (points bleus dans la Figure 1A) couvrant un angle de 90 °. La taille des éléments individuels de l&#39;ordre de 3 x 3 mm 2, ainsi que leur orientation et la bande passante de la fréquence (environ 2-6 MHz) garantissent collection de signal efficace d&#39;un volume centimétrique entourant le centre de la sphère (cube noir sur la figure 1A). Excitation optique de la région de formation d&#39;image est muni d&#39;un faisceau de fibres inséré à travers une cavité cylindrique centrale de la matrice, de sorte que la longueur d&#39;onde quelconque SUSCeptible d&#39;être transmise à travers le faisceau de fibres peut être utilisé pour l&#39;imagerie. Une image réelle de la matrice de transducteurs avec le faisceau de fibres optiques est représentée sur la figure 1B. L&#39;excitation efficace et la détection simultanée de signaux permet l&#39;imagerie des tissus profonds avec une seule photo excitation (une impulsion laser), de sorte que l&#39;imagerie en temps réel à une cadence déterminée par la fréquence de répétition des impulsions du laser est plus activé avec un Graphics- traitement-unité (GPU) la mise en œuvre de la procédure de reconstruction 17. Un boîtier cylindrique avec une membrane de polyéthylène transparent (figure 1C) est fixé à la matrice de transducteur pour enfermer un milieu liquide transmettant acoustique (de l&#39;eau). La membrane est en outre couplé au tissu au moyen d&#39;un gel acoustique. Une image de la sonde acousto-optique d&#39;être utilisé en mode de fonctionnement à main est illustré à la figure 1D. Le thr démontrédimensions imagerie acousto-optique à main ee combinée avec le temps réel la capacité de l&#39;imagerie fonctionnelle venir avec des avantages importants pour les diagnostics cliniques et un certain nombre d&#39;applications potentielles sont envisagées pour diverses indications, telles que la maladie vasculaire périphérique, du système lymphatique, le cancer du sein, des lésions cutanées, l&#39;inflammation ou de l&#39;arthrite 18. En outre, la capacité d&#39;imagerie rapide permet la visualisation d&#39;événements biologiques dynamiques avec la sonde disposé dans une position stationnaire. En combinaison avec l&#39;oscillateur paramétrique optique (OPO), la technologie laser rapide en longueur d&#39;onde d&#39;accord, cette approche permet une imagerie en temps réel de la biodistribution des agents photo-absorbant. Ainsi, de nouvelles possibilités peuvent également émerger dans de petites applications d&#39;imagerie animale, par exemple., Dans l&#39;étude de l&#39;hémodynamique des tissus, suivi in vivo dans des cellules, la visualisation de la pharmacocinétique, la perfusion des organes, ciblé imagerie moléculaire des tumeurs et le système cardiovasculaire, ou neuro-imagerie. <p class = &quot;jove_content&quot;&gt; Dans ce travail, nous présentons une description détaillée du protocole d&#39;imagerie expérimental pour fonctionner avec le réseau sphérique acousto-optique sonde et vitrine performances portatif dans plusieurs scénarios cliniques et petits typiques imagerie animale.

<table border="0" cellpadding="0" cellspacing="0" width="1663">
	<tbody>
		<tr height="21">
			<td height="21" style="height:21px;width:327px;">Name of Material/ Equipment</td>
			<td style="width:389px;">Company</td>
			<td style="width:171px;">Catalog Number</td>
			<td style="width:777px;">Comments/Description</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:327px;">Optical parametric oscillator (OPO)-based laser</td>
			<td style="width:389px;">Innolas Laser GmbH, Krailling (Germany)</td>
			<td style="width:171px;">custom-made</td>
			<td style="width:777px;">The laser provides laser pulses with a duration around 10ns and an energy up to 80mJ. 
			The wavelength is tunable between 680-950nm.</td>
		</tr>
		<tr height="45">
			<td height="45" style="height:45px;width:327px;">Spherical array of piezocomposite detectors</td>
			<td style="width:389px;">Imasonic SaS, Voray (France)</td>
			<td style="width:171px;">custom-made</td>
			<td style="width:777px;">The array consists of 256 piezoelectric sensors distributed on a spherical surface. 
			Each element has dimensions 3x3mm2, a central frequency of 4MHz and a bandwidth of 100%.&#160;</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:327px;">Data acquisition system (DAQ)</td>
			<td style="width:389px;">Falkenstein Mikrosysteme GmbH, Taufkirchen (Germany)</td>
			<td style="width:171px;">custom-made</td>
			<td style="width:777px;">The DAQ simultaneously acquires 256 signals at 40 megasamples per second and 2030 samples. 
			The input impedance is 1MW.</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:327px;">Fiber bundle</td>
			<td style="width:389px;">CeramOptec GmbH, Bonn (Germany)</td>
			<td style="width:171px;">custom-made</td>
			<td style="width:777px;">The bundle consists of 480 individual fibers randomly distributed in the input and output. 
			The numerical aperture of each individual fiber is 0.22.</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:327px;">Athymic Nude mouse</td>
			<td style="width:389px;">Harlan Laboratories (The Netherlands)</td>
			<td style="width:171px;">Athymic nude - Foxn1nu</td>
			<td style="width:777px;">The mouse was 8 weeks old (adult) at the time of the experiment. 
			The ethical protocol was approved by the Bavarian goverment (number 55.2.1.54-2632-102-11)</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:327px;">Bepanthen cream</td>
			<td style="width:389px;">Bayer AG (Germany)</td>
			<td style="width:171px;">N/A</td>
			<td>Vet ointment to protect the eyes during anesthesia</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:327px;">Data processing software</td>
			<td style="width:389px;">Matlab (Mathworks, Natick, MA, USA)</td>
			<td style="width:171px;">custom-made</td>
			<td style="width:777px;">The data processing software was devoped at our institute. 
			It allows reconstruction at each wavelength and multi-wavelength unmixing, as well as further data processing.</td>
		</tr>
		<tr height="24">
			<td height="24" style="height:24px;width:327px;">Water-enclosing part</td>
			<td style="width:389px;">N/A</td>
			<td style="width:171px;">custom-made</td>
			<td>This part contains the water that acts as an acoustic coupling medium between skin and transducer elements</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:327px;">Indocyanine green (ICG)</td>
			<td style="width:389px;">PULSION Medical Systems SE</td>
			<td style="width:171px;">N/A</td>
			<td style="width:777px;">ICG-PULSION (active ingredient: indocyanine green dye) is a drug used in cardiac, circulatory and micro-circulatory diagnostics, liver function diagnostics and ophthalmic angiography diagnostics.</td>
		</tr>
	</tbody>
</table>

universal hand-held three-dimensional optoacoustic imaging probe for deep tissue human angiography and functional preclinical studies in real time

The exclusive combination of high optical contrast and excellent spatial resolution makes optoacoustics (photoacoustics) ideal for simultaneously attaining anatomical, functional and molecular contrast in deep optically opaque tissues. While enormous potential has been recently demonstrated in the application of optoacoustics for small animal research, vast efforts have also been undertaken in translating this imaging technology into clinical practice. We present here a newly developed optoacoustic tomography approach capable of delivering high resolution and spectrally enriched volumetric images of tissue morphology and function in real time. A detailed description of the experimental protocol for operating with the imaging system in both hand-held and stationary modes is provided and showcased for different potential scenarios involving functional and molecular studies in murine models and humans. The possibility for real time visualization in three dimensions along with the versatile handheld design of the imaging probe make the newly developed approach unique among the pantheon of imaging modalities used in today&#8217;s preclinical research and clinical practice.

Les résultats représentatifs, ce qui démontre les capacités de la sonde volumétrique décrit à main acousto-optique, sont présentés dans cette section. Dans tous les cas, l&#39;influence de lumière sur la surface de la peau a été maintenue en dessous de la limite d&#39;exposition de la sécurité de 20 mJ / cm 2 19. La performance de la sonde en temps réel le suivi vasculaire périphérique humain est présenté à la figure 2. Au cours de cette expér...

Watch this Scientific Journal Video about Universal Hand-held Three-dimensional Optoacoustic Imaging Probe for Deep Tissue Human Angiography and Functional Preclinical Studies in Real Time at JoVE.com

Universal Hand-held Three-dimensional Optoacoustic Imaging Probe for Deep Tissue Human Angiography and Functional Preclinical Studies in Real Time

We provide herein a detailed description of the experimental protocol for imaging with a newly developed hand-held optoacoustic (photoacoustic) system for three-dimensional functional and molecular imaging in real time. The demonstrated powerful performance and versatility may define new application areas of the optoacoustic technology in preclinical research and clinical practice.

Universal à main en trois dimensions optoacoustique sonde d&#39;imagerie pour Deep Tissue angiographie humain et des études précliniques fonctionnels en temps réel

The exclusive combination of high optical contrast and excellent spatial resolution makes optoacoustics (photoacoustics) ideal for simultaneously ...

universal-hand-held-three-dimensional-optoacoustic-imaging-probe-for

Research

JoVE Journal

Bioengineering

10.6K vues.  Helmholtz Zentrum München. We provide herein a detailed description of the experimental protocol for imaging with a newly developed hand-held optoacoustic (photoacoustic) system for three-dimensional functional and molecular imaging in real time. The demonstrated powerful performance and versatility may define new application areas of the optoacoustic technology in preclinical research and clinical practice.

Vidéo: Universal Hand-held Three-dimensional Optoacoustic Imaging Probe for Deep Tissue Human Angiography and Functional Preclinical Studies in Real Time

Three-dimensional Optical-resolution Photoacoustic Microscopy

Optical microscopy, providing valuable insights at the cellular and organelle levels, has been widely recognized as an enabling biomedical technology. As the mainstays of in vivo three-dimensional (3-D) optical microscopy, single-/multi-photon fluorescence microscopy and optical coherence tomography (OCT) have demonstrated their extraordinary sensitivities to fluorescence and optical scattering contrasts, respectively. However, the optical absorption contrast of biological tissues, which encodes essential physiological/pathological information, has not yet been assessable. 
The emergence of biomedical photoacoustics has led to a new branch of optical microscopy optical-resolution photoacoustic microscopy (OR-PAM)1, where the optical irradiation is focused to the diffraction limit to achieve cellular1 or even subcellular2 level lateral resolution. As a valuable complement to existing optical microscopy technologies, OR-PAM brings in at least two novelties. First and most importantly, OR-PAM detects optical absorption contrasts with extraordinary sensitivity (i.e., 100%). Combining OR-PAM with fluorescence microscopy3 or with optical-scattering-based OCT4 (or with both) provides comprehensive optical properties of biological tissues. Second, OR-PAM encodes optical absorption into acoustic waves, in contrast to the pure optical processes in fluorescence microscopy and OCT, and provides background-free detection. The acoustic detection in OR-PAM mitigates the impacts of optical scattering on signal degradation and naturally eliminates possible interferences (i.e., crosstalks) between excitation and detection, which is a common problem in fluorescence microscopy due to the overlap between the excitation and fluorescence spectra. 
Unique for optical absorption imaging, OR-PAM has demonstrated broad biomedical applications since its invention, including, but not limited to, neurology5, 6, ophthalmology7, 8, vascular biology9, and dermatology10. In this video, we teach the system configuration and alignment of OR-PAM as well as the experimental procedures for in vivo functional microvascular imaging.

Optical-resolution photoacoustic microscopy (OR-PAM) is an emerging technology capable of imaging optical absorption contrasts in vivo with cellular resolution and sensitivity. Here, we provide a visualized instruction on the experimental protocols of OR-PAM, including system configuration, system alignment, typical in vivo experimental procedures, and functional imaging schemes.

Tridimensionnelle optique résolution microscopie photoacoustique

Optical microscopy, providing valuable insights at the cellular and organelle levels, has been widely recognized as an enabling biomedical technology. As ...

Recherche

Bio-ingénierie

Planar and Three-Dimensional Printing of Conductive Inks

Printed electronics rely on low-cost, large-area fabrication routes to create flexible or multidimensional electronic, optoelectronic, and biomedical devices1-3. In this paper, we focus on one- (1D), two- (2D), and three-dimensional (3D) printing of conductive metallic inks in the form of flexible, stretchable, and spanning microelectrodes.
Direct-write assembly4,5 is a 1-to-3D printing technique that enables the fabrication of features ranging from simple lines to complex structures by the deposition of concentrated inks through fine nozzles (~0.1 - 250 &mu;m). This printing method consists of a computer-controlled 3-axis translation stage, an ink reservoir and nozzle, and 10x telescopic lens for visualization. Unlike inkjet printing, a droplet-based process, direct-write assembly involves the extrusion of ink filaments either in- or out-of-plane. The printed filaments typically conform to the nozzle size. Hence, microscale features (&lt; 1 &mu;m) can be patterned and assembled into larger arrays and multidimensional architectures.
In this paper, we first synthesize a highly concentrated silver nanoparticle ink for planar and 3D printing via direct-write assembly. Next, a standard protocol for printing microelectrodes in multidimensional motifs is demonstrated. Finally, applications of printed microelectrodes for electrically small antennas, solar cells, and light-emitting diodes are highlighted.

Planar and three-dimensional printing of conductive metallic inks is described. Our approach provides new avenues for fabricating printed electronic, optoelectronic, and biomedical devices in unusual layouts at the microscale.

Impression Planar et en trois dimensions des encres conductrices

Printed electronics rely on low-cost, large-area fabrication routes to create flexible or multidimensional electronic, optoelectronic, and biomedical ...

A Novel Three-dimensional Flow Chamber Device to Study Chemokine-directed Extravasation of Cells Circulating under Physiological Flow Conditions

Extravasation of circulating cells from the bloodstream plays a central role in many physiological and pathophysiological processes, including stem cell homing and tumor metastasis. The three-dimensional flow chamber device (hereafter the 3D device) is a novel in vitro technology that recreates physiological shear stress and allows each step of the cell extravasation cascade to be quantified. The 3D device consists of an upper compartment in which the cells of interest circulate under shear stress, and a lower compartment of static wells that contain the chemoattractants of interest. The two compartments are separated by porous inserts coated with a monolayer of endothelial cells (EC). An optional second insert with microenvironmental cells of interest can be placed immediately beneath the EC layer. A gas exchange unit allows the optimal CO2 tension to be maintained and provides an access point to add or withdraw cells or compounds during the experiment. The test cells circulate in the upper compartment at the desired shear stress (flow rate) controlled by a peristaltic pump. At the end of the experiment, the circulating and migrated cells are collected for further analyses. The 3D device can be used to examine cell rolling on and adhesion to EC under shear stress, transmigration in response to chemokine gradients, resistance to shear stress, cluster formation, and cell survival. In addition, the optional second insert allows the effects of crosstalk between EC and microenvironmental cells to be examined. The translational applications of the 3D device include testing of drug candidates that target cell migration and predicting the in vivo behavior of cells after intravenous injection. Thus, the novel 3D device is a versatile and inexpensive tool to study the molecular mechanisms that mediate cellular extravasation.

The three-dimensional flow chamber device is a novel in vitro technology for the quantitative and step-wise evaluation of the extravasation cascade of cells circulating under conditions of physiological shear stress. The device therefore fills a critical need for basic, preclinical, and clinical studies of cell migration.

A Novel tridimensionnelle chambre d&#39;écoulement périphérique pour étudier chimiokine dirigée extravasation des cellules circulantes des conditions d&#39;écoulement physiologiques

Extravasation of circulating cells from the bloodstream plays a central role in many physiological and pathophysiological processes, including stem cell ...

Human Cartilage Tissue Fabrication Using Three-dimensional Inkjet Printing Technology

Bioprinting, which is based on thermal inkjet printing, is one of the most attractive enabling technologies in the field of tissue engineering and regenerative medicine. With digital control&#160;cells, scaffolds, and growth factors can be precisely deposited to the desired two-dimensional (2D) and three-dimensional (3D) locations rapidly. Therefore, this technology is an ideal approach to fabricate tissues mimicking their native anatomic structures. In order to engineer cartilage with native zonal organization, extracellular matrix composition (ECM), and mechanical properties, we developed a bioprinting platform using a commercial inkjet printer with simultaneous photopolymerization capable for 3D cartilage tissue engineering. Human chondrocytes suspended in poly(ethylene glycol) diacrylate (PEGDA) were printed for 3D neocartilage construction via layer-by-layer assembly. The printed cells were fixed at their original deposited positions, supported by the surrounding scaffold in simultaneous photopolymerization. The mechanical properties of the printed tissue were similar to the native cartilage. Compared to conventional tissue fabrication, which requires longer UV exposure, the viability of the printed cells with simultaneous photopolymerization was significantly higher. Printed neocartilage demonstrated excellent glycosaminoglycan (GAG) and collagen type II production, which was consistent with gene expression. Therefore, this platform is ideal for accurate cell distribution and arrangement for anatomic tissue engineering.

The methods described in this paper show how to convert a commercial inkjet printer into a bioprinter with simultaneous UV polymerization. The printer is capable of constructing 3D tissue structure with cells and biomaterials. The study demonstrated here constructed a 3D neocartilage.

Fabrication de tissus de cartilage humain en utilisant trois dimensions jet d&#39;encre Technologie d&#39;impression

Bioprinting, which is based on thermal inkjet printing, is one of the most attractive enabling technologies in the field of tissue engineering and ...

Generation of a Three-dimensional Full Thickness Skin Equivalent and Automated Wounding

In vitro models are a cost effective and ethical alternative to study cutaneous wound healing processes. Moreover, by using human cells, these models reflect the human wound situation better than animal models. Although two-dimensional models are widely used to investigate processes such as cellular migration and proliferation, models that are more complex are required to gain a deeper knowledge about wound healing. Besides a suitable model system, the generation of precise and reproducible wounds is crucial to ensure comparable results between different test runs. In this study, the generation of a three-dimensional full thickness skin equivalent to study wound healing is shown. The dermal part of the models is comprised of human dermal fibroblast embedded in a rat-tail collagen type I hydrogel. Following the inoculation with human epidermal keratinocytes and consequent culture at the air-liquid interface, a multilayered epidermis is formed on top of the models. To study the wound healing process, we additionally developed an automated wounding device, which generates standardized wounds in a sterile atmosphere.

The goal of this protocol is to build up a three-dimensional full thickness skin equivalent, which resembles natural skin. With a specifically constructed automated wounding device, precise and reproducible wounds can be generated under maintenance of sterility.

Génération d&#39;une épaisseur en trois dimensions équivalent de peau et automatisé Blessant

In vitro models are a cost effective and ethical alternative to study cutaneous wound healing processes. Moreover, by using human cells, these models ...

Development of a Multicellular Three-dimensional Organotypic Model of the Human Intestinal Mucosa Grown Under Microgravity

Because cells growing in a three-dimensional (3-D) environment have the potential to bridge many gaps of cell cultivation in 2-D environments (e.g., flasks or dishes). In fact, it is widely recognized that cells grown in flasks or dishes tend to de-differentiate and lose specialized features of the tissues from which they were derived. Currently, there are mainly two types of 3-D culture systems where the cells are seeded into scaffolds mimicking the native extracellular matrix (ECM): (a) static models and (b) models using bioreactors. The first breakthrough was the static 3-D models. 3-D models using bioreactors such as the rotating-wall-vessel (RWV) bioreactors are a more recent development. The original concept of the RWV bioreactors was developed at NASA&#39;s Johnson Space Center in the early 1990s and is believed to overcome the limitations of static models such as the development of hypoxic, necrotic cores. The RWV bioreactors might circumvent this problem by providing fluid dynamics that allow the efficient diffusion of nutrients and oxygen. These bioreactors consist of a rotator base that serves to support and rotate two different formats of culture vessels that differ by their aeration source type: (1) Slow Turning Lateral Vessels (STLVs) with a co-axial oxygenator in the center, or (2) High Aspect Ratio Vessels (HARVs) with oxygenation via a flat, silicone rubber gas transfer membrane. These vessels allow efficient gas transfer while avoiding bubble formation and consequent turbulence. These conditions result in laminar flow and minimal shear force that models reduced gravity (microgravity) inside the culture vessel. Here we describe the development of a multicellular 3-D organotypic model of the human intestinal mucosa composed of an intestinal epithelial cell line and primary human lymphocytes, endothelial cells and fibroblasts cultured under microgravity provided by the RWV bioreactor.

Cells growing in a three-dimensional (3-D) environment represent a marked improvement over cell cultivation in 2-D environments (e.g., flasks or dishes). Here we describe the development of a multicellular 3-D organotypic model of the human intestinal mucosa cultured under microgravity provided by rotating-wall-vessel (RWV) bioreactors.

Développement d&#39;un modèle organotypique multicellulaire en trois dimensions de la muqueuse intestinale humaine Grown Sous microgravité

Because cells growing in a three-dimensional (3-D) environment have the potential to bridge many gaps of cell cultivation in 2-D environments (e.g., ...

Three-dimensional Tissue Engineered Aligned Astrocyte Networks to Recapitulate Developmental Mechanisms and Facilitate Nervous System Regeneration

Neurotrauma and neurodegenerative disease often result in lasting neurological deficits due to the limited capacity of the central nervous system (CNS) to replace lost neurons and regenerate axonal pathways. However, during nervous system development, neuronal migration and axonal extension often occur along pathways formed by other cells, referred to as &#34;living scaffolds&#34;. Seeking to emulate these mechanisms and to design a strategy that circumvents the inhibitory environment of the CNS, this manuscript presents a protocol to fabricate tissue engineered astrocyte-based &#34;living scaffolds&#34;. To create these constructs, we employed a novel biomaterial encasement scheme to induce astrocytes to self-assemble into dense three-dimensional bundles of bipolar longitudinally-aligned somata and processes. First, hollow hydrogel micro-columns were assembled, and the inner lumen was coated with collagen extracellular-matrix. Dissociated cerebral cortical astrocytes were then delivered into the lumen of the cylindrical micro-column and, at a critical inner diameter of &#60;350 &#181;m, spontaneously self-aligned and contracted to produce long fiber-like cables consisting of dense bundles of astrocyte processes and collagen fibrils measuring &#60;150 &#181;m in diameter yet extending several cm in length. These engineered living scaffolds exhibited &#62;97% cell viability and were virtually exclusively comprised of astrocytes expressing a combination of the intermediate filament proteins glial-fibrillary acidic protein (GFAP), vimentin, and nestin. These aligned astrocyte networks were found to provide a permissive substrate for neuronal attachment and aligned neurite extension. Moreover, these constructs maintain integrity and alignment when extracted from the hydrogel encasement, making them suitable for CNS implantation. These preformed constructs structurally emulate key cytoarchitectural elements of naturally occurring glial-based &#34;living scaffolds&#34; in vivo. As such, these engineered living scaffolds may serve as test-beds to study neurodevelopmental mechanisms in vitro or facilitate neuroregeneration by directing neuronal migration and/or axonal pathfinding following CNS degeneration in vivo.

We showcase the development of self-assembled, three-dimensional bundles of longitudinally aligned astrocytic somata and processes within a novel biomaterial encasement. These engineered &#34;living scaffolds&#34;, exhibiting micron-scale diameter yet extending centimeters in length, may serve as test-beds to study neurodevelopmental mechanisms or facilitate neuroregeneration by directing neuronal migration and/or axonal pathfinding.

Dimensions des tissus conçus alignement Astrocyte réseaux pour récapituler les mécanismes du développement et de faciliter la régénération du système nerveux

Neurotrauma and neurodegenerative disease often result in lasting neurological deficits due to the limited capacity of the central nervous system (CNS) to ...

Melt Electrospinning Writing of Three-dimensional Poly(&#949;-caprolactone) Scaffolds with Controllable Morphologies for Tissue Engineering Applications

This tutorial reflects on the fundamental principles and guidelines for electrospinning writing with polymer melts, an additive manufacturing technology with great potential for biomedical applications. The technique facilitates the direct deposition of biocompatible polymer fibers to fabricate well-ordered scaffolds in the sub-micron to micro scale range. The establishment of a stable, viscoelastic, polymer jet between a spinneret and a collector is achieved using an applied voltage and can be direct-written. A significant benefit of a typical porous scaffold is a high surface-to-volume ratio which provides increased effective adhesion sites for cell attachment and growth. Controlling the printing process by fine-tuning the system parameters enables high reproducibility in the quality of the printed scaffolds. It also provides a flexible manufacturing platform for users to tailor the morphological structures of the scaffolds to their specific requirements. For this purpose, we present a protocol to obtain different fiber diameters using melt electrospinning writing (MEW) with a guided amendment of the parameters, including flow rate, voltage and collection speed. Furthermore, we demonstrate how to optimize the jet, discuss often experienced technical challenges, explain troubleshooting techniques and showcase a wide range of printable scaffold architectures.

Melt Electrospinning Writing of Three-dimensional Poly(&amp;#949;-caprolactone) Scaffolds with Controllable Morphologies for Tissue Engineering Applications

This protocol serves as a comprehensive guideline to fabricate scaffolds via electrospinning with polymer melts in a direct writing mode. We systematically outline the process and define the appropriate parameter settings for achieving targeted scaffold architectures.

Faire fondre électrofilage écrit d’échafaudages Poly(ε-caprolactone) en trois dimensions avec des Morphologies contrôlables pour les Applications de génie tissulaire

This tutorial reflects on the fundamental principles and guidelines for electrospinning writing with polymer melts, an additive manufacturing technology ...

Hand-held Clinical Photoacoustic Imaging System for Real-time Non-invasive Small Animal Imaging

Translation of photoacoustic imaging into the clinic is a major challenge. Handheld real-time clinical photoacoustic imaging systems are very rare. Here, we report a combined photoacoustic and clinical ultrasound imaging system by integrating an ultrasound probe with light delivery for small animal imaging. We demonstrate this by showing sentinel lymph node imaging in small animals along with minimally invasive real-time needle guidance. A clinical ultrasound platform with access to raw channel data allows the integration of photoacoustic imaging leading to a handheld real-time clinical photoacoustic imaging system. Methylene blue was used for sentinel lymph node imaging at 675 nm wavelength. Additionally, needle guidance with dual modal ultrasound and photoacoustic imaging was shown using the imaging system. Depth imaging of up to 1.5 cm was demonstrated with a 10 Hz laser at a photoacoustic imaging frame rate of 5 frames per second.

A clinical handheld photoacoustic imaging system will be demonstrated for real-time non-invasive small animal imaging.

À main photoacoustique clinique, système d’imagerie pour l’imagerie animale petit Non invasif en temps réel

Translation of photoacoustic imaging into the clinic is a major challenge. Handheld real-time clinical photoacoustic imaging systems are very rare. Here, ...

Culturing Mammalian Cells in Three-dimensional Peptide Scaffolds

A useful technique for culturing cells in a self-assembling nanofiber three-dimensional (3D) scaffold is described. This culture system recreates an environment that closely mimics the structural features of non-polarized tissue. Furthermore, the particular intrinsic nanofiber structure of the scaffold makes it transparent to visual light, which allows for easy visualization of the sample under microscopy. This advantage was largely used to study cell migration, organization, proliferation, and differentiation and thus any development of their particular cellular function by staining with specific dyes or probes. Furthermore, in this work, we describe the good performance of this system to easily study the redifferentiation of expanded human articular chondrocytes into cartilaginous tissue. Cells were encapsulated into self-assembling peptide scaffolds and cultured under specific conditions to promote chondrogenesis. Three-dimensional cultures showed good viability during the 4 weeks of the experiment. As expected, samples cultured with chondrogenic inducers (compared to non-induced controls) stained strongly positive for toluidine blue (which stains glycosaminoglycans (GAGs) that are highly present in cartilage extracellular matrix) and expressed specific molecular markers, including collagen type I, II and X, according to Western Blot analysis. This protocol is easy to perform and can be used at research laboratories, industries and for educational purposes in laboratory courses.

Here, we present a protocol to obtain 3D-culture systems in self-assembling peptide scaffolds to promote the differentiation of dedifferentiated human articular chondrocytes into cartilage-like tissue.

Culture de cellules mammifères dans les échafaudages de Peptide en trois dimensions

A useful technique for culturing cells in a self-assembling nanofiber three-dimensional (3D) scaffold is described. This culture system recreates an ...