Ce travail a été soutenu par la Princess Margaret Hospital Foundation, la National Science et génie du Canada, l&#39;Institut canadien de recherche en santé, la Fondation canadienne pour l&#39;innovation, et le Joey et Toby Tanenbaum / Ball Chair brésilien en recherche sur le cancer de la prostate, un subvention de la Fondation SUNY recherche et assistance au démarrage de l&#39;Université de Buffalo.

Une. Synthèse du lipide-Pyropheophorbide <ol><li> Moissonneuse 200 mmol pyropheophorbide (préparé à partir d&#39;algues Spirulina Pacifica, comme décrit précédemment,... Zheng et al, Bioconj Chem, 2002, 13 -392) 4, 98,7 mg 16:00 lysophophatidylcholine (1-palmitoyl-2-hydroxy-sn-glycéro -3-phosphocholine, Avanti Polar Lipids # 855675), 76,3 mg d&#39;EDC, 48,7 mg de DMAP (4 - (diméthylamino) pyridine) dans 5 ml amylène stabilisé chloroforme pour un ratio de 1:1:2:2 Pyro: lipides: EDC : DMPA. </li><li> On agite le mélange réactionnel à la température ambiante sous argon pendant 24 heures. On évapore le chloroforme sous pression réduite en utilisant un évaporateur rotatif avec un chauffage à 40 ° C. Le mélange d&#39;isomères de pyro-lipide (pyro attachés à la position sn-1 ou sn-2) est produite. </li><li> mélange d&#39;isomères de pyro-lipide Remettre en suspension dans du CaCl2 1 mM, Tris 50 mM (pH 8), 0,5% de Triton X-100 et 10% de méthanol à la concentration de 5 mg / ml. Ajouter phospholipase A2 (PLA2) de venin d&#39;abeille à0,1 mg / ml et la concentration de la solution incuber à 37 ° C pendant 24 heures pour digérer sn-1 et pyro-lipides pour produire le sn-2 pyro-lipide isomère pur. </li><li> Ajouter 2 volumes supplémentaires de chloroforme et 1,25 volumes de méthanol, de l&#39;extrait, puis éliminer le solvant en utilisant un évaporateur rotatif sous pression réduite à 40 ° C. Remettre en suspension le produit de clivage dans 1% de MeOH dans du DCM et on purifie sur une colonne de silice de faible diol pour la pyro-lipide isomère pur. </li></ol> 2. Préparation de porphysomes <ol><li> Dissoudre pyropheophorbide-lipide, distéaroyl-sn-glycéro-3-phosphoéthanolamine-N-méthoxy (polyéthylène glycol) (PEG-2000-PE) et le taux de cholestérol dans du chloroforme séparément, et l&#39;enregistrement des concentrations respectivement. </li><li> Préparer porphysome film lipidique dans un tube à essai de borosilicate de 12x75 mm avec un poids total en lipides de 5 mg / tuyau en combinant 65% molaire lipide-porphyrine, 5% en moles de PEG-2000-PE et 30% molaire de cholestérol. </li><li> Sécher les films lipidiques sous un courant d&#39;nitrogaz gen et laissez-les encore séchés sous vide pendant 1 heure. Stocker les films lipidiques à -20 ° C sous atmosphère d&#39;argon jusqu&#39;à ce que l&#39;hydratation et l&#39;extrusion ultérieure. </li><li> Ajouter 1 ml de solution saline de phosphate tamponnée (PBS, NaCl 150 mM, phosphate 10 mM, pH 7,4) à chaque tube de film lipidique. Congeler le tube à essai dans l&#39;azote liquide jusqu&#39;à ce que complètement gelé, dégeler le tube 65 ° C bain d&#39;eau jusqu&#39;à ce que l&#39;échantillon a tous fondu. Vortexer le tube pendant un total de 30 sec (3 x 10 sec à chacune, avec 5 secondes de réchauffage à 65 ° C bain d&#39;eau entre chaque vortex). </li><li> Répétez le gel-dégel pendant 5 cycles ou plus sur nécessaire jusqu&#39;à ce qu&#39;il n&#39;y grands agrégats peuvent être observées dans la solution hydratée. Laisser la suspension de porphysome sur la glace lorsque vous avez terminé. </li><li> Assembler l&#39;extrudeuse à haute pression (10 ml) avec deux empilés 100 filtres nm en polycarbonate installés po Connect l&#39;extrudeuse de thermobarrel à un circulateur thermostaté 65 ° C bain à circulation d&#39;eau, et connecter le système de l&#39;extrudeuse à un réservoir d&#39;azote pour haute pressionque l&#39;alimentation. </li><li> Transférer la suspension de porphysome à la chambre de l&#39;extrudeuse en utilisant une pipette de transfert en verre de 9 pouces. Régler la pression en tournant la molette du régulateur de réservoir d&#39;azote jusqu&#39;à obtenir une lecture entre 200 et 300 psig on observe que la pression d&#39;extrusion initiale. Ouvrir la vanne du régulateur de pression graduellement jusqu&#39;à ce que le débit d&#39;porphysome suffisant est atteint, soit un maximum de 800 psig (5440 kPa) de pression est atteinte. </li><li> Après le premier passage d&#39;extrusion est terminée, fermer la soupape de commande de pression et de libérer la pression de l&#39;extrudeuse par l&#39;ouverture de la soupape de décharge de pression sur la partie supérieure de l&#39;extrudeuse. Remplir l&#39;extrudeuse avec la solution de porphysome fraîchement extrudée, et répéter l&#39;extrusion d&#39;au moins 10 fois pour s&#39;assurer que la solution finale est extrudé homogène. </li><li> Déterminer la concentration de la solution finale de porphysome en mesurant l&#39;absorption d&#39;un échantillon dilué dans du méthanol par spectromètre UV-Vis. Calculer la concentration en utilisant l&#39;esprit de l&#39;équation suivanteh le coefficient d&#39;extinction de 97000 M -1 cm -1 à 410 nm pour pyropheophorbide-lipide. </li></ol><img alt="Equation 1" src="/files/ftp_upload/50536/50536eq1.jpg" /><ol start="10"><li> Mesurer la distribution de la taille finale du porphysomes utilisant un Malvern Zetasizer ZS90 par diffusion de lumière dynamique. Diluer la solution de porphysome dans du PBS et effectuer trois mesures avec 15 points chacun pour les moyennes des résultats. </li><li> Déterminer l&#39;extinction de fluorescence de porphysome en comparant l&#39;émission de fluorescence de porphysome trempé (1 mM dans du PBS) et porphysome non éteint (1 mM dans 1% de Triton X-100) en utilisant Spectrofluorometer. Prendre les spectres avant et après le détergent (Tirton X-100) est ajouté, et à normaliser le signal de lecture de la fluorescence maximale. Régler l&#39;excitation à 410 nm, et l&#39;émission de longueur d&#39;onde de 600-800 nm être. </li><li> Conserver la solution de porphysomes sous argon à 4 ° C jusqu&#39;à leur utilisation future, et protéger de la lumière par l&#39;aluminium fhuile. </li></ol> 3. Préparation des animaux modèle de xénogreffe <ol><li> Culture des cellules KB en RPMI-1640 (avec 10% de FBS), et maintenir les cellules dans 5% de CO 2 et de l&#39;atmosphère hydratée à 37 ° C. </li><li> Cellules récolte KB utilisant la technique de culture cellulaire standard. Gardez la suspension cellulaire (20 000 cellules / ml) dans un tampon phosphate salin (PBS) sur la glace pour le stockage à court terme jusqu&#39;à ce moment de l&#39;injection. </li><li> Préparer un KB xénogreffes sur souris nude. Souris nude sont achetés auprès de Charles River Laboratories quand ils sont 20 g (Catégorie # Crl: NU (RCN)-Foxn1 nu, 6-7 semaines-vieux), compte tenu de régime standard et la literie (Harlan Laboratories; Régime alimentaire: irradiés LM-485, # 7912; Literie: literie Corn ¼ &quot;, # 7097). Stériliser tous les outils de manipulation des animaux (seringues, des pinces, et pad animale) avant les expériences. Anesthésier les souris avec 2% d&#39;isoflurane v / v dans 100% gaz porteur d&#39;oxygène. Tous les protocoles d&#39;animaux sont approuvés par l&#39;institution (University Health Network dans ce cas) le soin des animaux et l&#39;utilisation comité. </li><li> Injecter les cellules tumorales en suspension dans du PBS par voie sous cutanée à la flanc droit de souris. Les souris sont prêtes à être utilisées après 10 jours lorsque la tumeur atteint un diamètre de 4-5 mm et une épaisseur de 3.2 mm. </li></ol> 4. In vivo thérapie photothermique <ol><li> Injecter porphysomes (contenant 750 nmol pyro à 200 volume de pi) par voie intraveineuse via la veine caudale. Effectuer irradiation laser 24 h post-injection de porphysomes. </li><li> Dans cette section, mener toutes les procédures dans une salle de sécurité laser clos. Porter des lunettes de sécurité laser lors de l&#39;irradiation laser. </li><li> Mettre en place l&#39;équipement d&#39;irradiation laser. Maintenir la fibre de laser et d&#39;un diffuseur par une pince sur un support. Réglez la puissance du laser à faible niveau et activer le laser. Réglez la hauteur de la fibre laser pour atteindre une zone d&#39;irradiation de 9 mm de diamètre (DPSS Laser, LaserGlow Technologies, Toronto, Canada), de calibrer la puissance du laser à 750 mW (1,18 W / cm 2) à l&#39;aide apeur mètres puis éteindre le laser à préparer les animaux. </li><li> Anesthésier les animaux avec 2% d&#39;isoflurane v / v dans 100% gaz porteur d&#39;oxygène par le nez cône pas évident de balayage de l&#39;isoflurane, un danger de risque pour la santé humaine). Assurez-vous que l&#39;animal est anesthésié assez profond en effectuant une pincée de pied avec des pinces ou des doigts à pattes arrière de l&#39;animal. Les animaux sont prêts pour l&#39;irradiation laser quand ils ne répondent pas à cette procédure. </li><li> Surveiller la température de la tumeur par une caméra infrarouge thermique (Mikroshot, LumaSense Technologies). Placez l&#39;appareil à proximité de la tumeur, se concentrer bien et prendre une image pré PTT ainsi que d&#39;une image en lumière blanche régulière avant PTT irradiation commence. </li><li> Allumez le laser de nouveau, et d&#39;augmenter la puissance lumineuse de 750 mW. Placez la tumeur dans le centre d&#39;un faisceau laser pour obtenir la tumeur entièrement couvert, et commencer à chronométrer l&#39;irradiation. Prendre l&#39;image de la température de la tumeur toutes les 5 s pour un traitement au laser d&#39;une minute. Éteignez le laser à 1 min. Stop l&#39;alimentation en gaz de 2% d&#39;isoflurane, attendez jusqu&#39;à ce que l&#39;animal se de l&#39;anesthésie et le remettre à la cage. </li><li> Gardez la souris à l&#39;animalerie de la normothermie sous 12h12 h cycle d&#39;obscurité et de la lumière selon des protocoles animales approuvées par l&#39;University Health Network. Injecter buprénorphine (0,05 mg / kg, voie sous-cutanée deux fois par jour pendant une semaine après l&#39;irradiation laser pour réduire la douleur causée par la cicatrice. Tous les deux jours, mesurer le diamètre de la tumeur à l&#39;aide d&#39;un pied et prendre une photo. Calculer le volume de la tumeur à l&#39;aide du l&#39;équation suivante: V = π / 6 · une · b 2, où a est le grand diamètre, et b le court. </li><li> Sacrifier les animaux en les mettant en CO 2 quand la chambre de grand diamètre de la tumeur atteint 10 mm, car il est le point final défini dans notre laboratoire. Effectuer dislocation cervicale après le traitement du CO2. Tracer la courbe de survie. </li></ol>

<ol>
	<li>Lovell, J. F., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Porphysome+nanovesicles+generated+by+porphyrin+bilayers+for+use+as+multimodal+biophotonic+contrast+agents.">Porphysome nanovesicles generated by porphyrin bilayers for use as multimodal biophotonic contrast agents.</a> Nat. Mater. 10, 324-332 (2011).</li><li>Lovell, J. F., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Enzymatic+regioselection+for+the+synthesis+and+biodegradation+of+porphysome+nanovesicles.">Enzymatic regioselection for the synthesis and biodegradation of porphysome nanovesicles.</a> Angew Chem Int Ed Engl. 51, 2429-2433 (2012).</li><li>Liu, T. W., MacDonald, T. D., Shi, J., Wilson, B. C., Zheng, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Intrinsically+Copper-64-Labeled+Organic+Nanoparticles+as+Radiotracers.">Intrinsically Copper-64-Labeled Organic Nanoparticles as Radiotracers.</a> Angewandte Chemie. , (2012).</li><li>Zheng, G., 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=Low-density+lipoprotein+reconstituted+by+pyropheophorbide+cholesteryl+oleate+as+target-specific+photosensitizer.">Low-density lipoprotein reconstituted by pyropheophorbide cholesteryl oleate as target-specific photosensitizer.</a> Bioconjug Chem. 13, 392-396 (2002).</li><li>Chen, B., Pogue, B. W., Hasan, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Liposomal+delivery+of+photosensitising+agents.">Liposomal delivery of photosensitising agents.</a> Expert Opin Drug Deliv. 2, 477-487 (2005).</li><li>Jin, C. S., Zheng, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Liposomal+nanostructures+for+photosensitizer+delivery.">Liposomal nanostructures for photosensitizer delivery.</a> Lasers Surg Med. 43, 734-748 (2011).</li><li>Olson, F., Hunt, C. A., Szoka, F. C., Vail, W. J., Papahadjopoulos, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Preparation+of+liposomes+of+defined+size+distribution+by+extrusion+through+polycarbonate+membranes.">Preparation of liposomes of defined size distribution by extrusion through polycarbonate membranes.</a> Biochim Biophys Acta. 557, 9-23 (1979).</li><li>Berger, N., Sachse, A., Bender, J., Schubert, R., Brandl, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Filter+extrusion+of+liposomes+using+different+devices:+comparison+of+liposome+size,+encapsulation+efficiency,+and+process+characteristics.">Filter extrusion of liposomes using different devices: comparison of liposome size, encapsulation efficiency, and process characteristics.</a> Int J Pharm. 223, 55-68 (2001).</li><li>Jesorka, A., Orwar, O. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Liposomes:+technologies+and+analytical+applications.">Liposomes: technologies and analytical applications.</a> Annu Rev Anal Chem (Palo Alto Calif). 1, 801-832 (2008).</li><li>Lapinski, M. M., Castro-Forero, A., Greiner, A. J., Ofoli, R. Y., Blanchard, G. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Comparison+of+liposomes+formed+by+sonication+and+extrusion:+rotational+and+translational+diffusion+of+an+embedded+chromophore.">Comparison of liposomes formed by sonication and extrusion: rotational and translational diffusion of an embedded chromophore.</a> Langmuir. 23, 11677-11683 (2007).</li><li>Liu, D., Huang, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Size+Homogeneity+of+a+Liposome+Preparation+is+Crucial+for+Liposome+Biodistribution+In+Vivo.">Size Homogeneity of a Liposome Preparation is Crucial for Liposome Biodistribution In Vivo.</a> Journal of Liposome Research. 2, 57-66 (1992).</li><li>Zhang, Y., Lovell, J. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Porphyrins+as+Theranostic+Agents+from+Prehistoric+to+Modern+Times.">Porphyrins as Theranostic Agents from Prehistoric to Modern Times.</a> Theranostics. 2, 905-915 (2012).</li></ol>

Nous n&#39;avons rien à communiquer.

Dans le développement des technologies de délivrance de médicaments, des nanoparticules multifonctionnelles sont actuellement en cours de larges investigations pour un suivi précis du véhicule de délivrance de médicament, tout en maintenant une charge de médicament élevée. liposomes de porphyrines chargé ont été développés pour de meilleures propriétés pharmacocinétiques et une prestation plus efficace que l&#39;administration directe de la porphyrine, mais des obstacles existent, y compris le montant ...

Porphysomes sont de nouveaux nanovésicules multifonctions que nous avons récemment développées qui sont capables de l&#39;imagerie et la thérapie 1 multimodal. Ils sont formés à partir des bicouches de porphyrine auto-assemblées et contiennent une densité extrêmement élevée de la porphyrine (plus de 83, 000/porphysome particules), qui génère des résultats et le coefficient de fluorescence à auto-extinction de la structure dépendante unique de grande extinction. Porphysomes ont de bonnes vivo pharmacocinétiques et de biodistribution propriétés dans: ils présentent une demi-vie dans le sang de 12 h après l&#39;administration systématique et passivement s&#39;accumulent dans les tumeurs de xénogreffe de 7,5% ID / g à 24 h post-injection 2. Leur structure unique et propriétés physico-chimiques font porphysome un bon candidat pour l&#39;imagerie multimodale et la thérapie guidée par l&#39;image. Tout d&#39;abord, contenant la porphyrine, porphysomes sont intrinsèquement apte à l&#39;imagerie de fluorescence des tumeurs lors de l&#39;accumulation d&#39;une tumeur.En outre, chaque porphyrine comporte un site de radio-isotopes stables de chelation, par conséquent, peuvent être facilement porphysomes marquées avec des radio-isotopes tels que le 64Cu pour l&#39;imagerie PET 3. En outre, l&#39;énergie de la lumière absorbée est dissipée thermiquement sous laser exposition d&#39;irradiation lors porphysome structure est intacte, de sorte porphysomes présentent également l&#39;imagerie photo-acoustique unique et capacités PTT. Il a été montré que 24 h après l&#39;injection intraveineuse de porphysomes, l&#39;irradiation laser de la tumeur au-porphysome accumulé induit une augmentation rapide de la température et de l&#39;ablation de la tumeur photothermique forte. Cela démontre que porphysomes sont efficaces exhausteurs de photothermique avec coefficient d&#39;extinction aussi élevé que les nanoparticules d&#39;or (AuNPs) 1. D&#39;autre part, en comparaison avec d&#39;autres agents photothermiques inorganiques, y compris AuNPs, porphysomes montrent un avantage exceptionnel en matière de biosécurité en raison de leur nature organique. Porphysomes sont enzymatique biodégradable et induisent toxicit aigu minimaly chez la souris avec des doses intraveineuses allant jusqu&#39;à 1000 mg / kg 1. En outre, même à des liposomes, la grande base aqueuse de porphysomes pourrait être passivement ou activement chargé avec des agents thérapeutiques ou d&#39;imagerie. Les propriétés optiques et de la biocompatibilité des porphysomes démontrent le potentiel multimodal de nanoparticules organiques pour l&#39;imagerie et la thérapie biophotonique. Dans cet article, nous présentons la méthode de synthèse de conjugués pyropheophorbide-lipidique, la fabrication et la méthode de caractérisation de porphysomes utilisant extrusion à haute pression. In vivo PTT sur des souris est effectuée ainsi à démontrer l&#39;efficacité de porphysome activé PTT dans la tumeur traitement à l&#39;aide d&#39;un modèle de tumeur de xénogreffe sous-cutanée.

<table border="1">
	<tbody>
		<tr>
			<td>Name of Equipment</td>
			<td>Company</td>
			<td>Catalog Number</td>
			<td>Comments (optional)</td>
		</tr>
		<tr>
			<td>Rotary evaporator</td>
			<td>Thermo-Savant</td>
			<td>SPD131DDA</td>
			<td>&nbsp;</td>
		</tr>
		<tr>
			<td>Votex</td>
			<td>Scientific Industries</td>
			<td>SI-0236</td>
			<td>Model number: G560</td>
		</tr>
		<tr>
			<td>High pressure extruder</td>
			<td>LIPEX, Northern Lipids Inc.</td>
			<td>T.001</td>
			<td>10 ml Thermobarrel Extruder</td>
		</tr>
		<tr>
			<td>Heated bath circulators (Thermostatted circulator)</td>
			<td>Thermo SCIENTIFIC</td>
			<td>SC100-S5P</td>
			<td>&nbsp;</td>
		</tr>
		<tr>
			<td>Polycarbonate filters</td>
			<td>Avanti Polar Lipids</td>
			<td>610005</td>
			<td>Pore size 100 nm, and membrane diameter 19 mm</td>
		</tr>
		<tr>
			<td>Zetasizer</td>
			<td>Malvern Instruments</td>
			<td>ZS90</td>
			<td>&nbsp;</td>
		</tr>
		<tr>
			<td>UV/Visible Spectrophototmeter</td>
			<td>Varian Australia</td>
			<td>Cary 50 Bio UV/ Visible Spectrophotometer</td>
			<td>&nbsp;</td>
		</tr>
		<tr>
			<td>Spectrofluorometer</td>
			<td>HORIBA Scientific</td>
			<td>FluoroMax-4</td>
			<td>&nbsp;</td>
		</tr>
		<tr>
			<td>Transmission Electron Microscopy (TEM)</td>
			<td>Hitachi</td>
			<td>H-7000</td>
			<td>&nbsp;</td>
		</tr>
		<tr>
			<td>Cell culture incubator</td>
			<td>SANYO</td>
			<td>MCO-18AIC</td>
			<td>&nbsp;</td>
		</tr>
		<tr>
			<td>Powermeter</td>
			<td>Thorlabs</td>
			<td>PM100D</td>
			<td>with sensor S142C</td>
		</tr>
		<tr>
			<td>671nm Laser</td>
			<td>LaserGlow Technologies</td>
			<td>LRS-0671_PFN-02000-05</td>
			<td>S/N:10097270</td>
		</tr>
		<tr>
			<td>Infrared Thermometer</td>
			<td>Mikroshot, LUMASENSE Technologies</td>
			<td>9102409</td>
			<td>&nbsp;</td>
		</tr>
		<tr>
			<td>Laser protective eye goggles</td>
			<td>LaserGlow Technologies</td>
			<td>AGF6602XX</td>
			<td>Optical Density: 1.5+ at 630-700 nm</td>
		</tr>
	</tbody>
</table>

one minute, sub-one-watt photothermal tumor ablation using porphysomes, intrinsic multifunctional nanovesicles

Nous avons récemment développé porphysomes comme nanovésicules intrinsèquement multifonctionnels. Un photosensibilisateur, pyropheophorbide α, a été conjugué à un phospholipide et l&#39;auto-assemblage de vésicules sphériques liposomes comme. En raison de la très forte densité de porphyrine dans la bicouche lipidique de porphyrine, porphysomes générées grands coefficients d&#39;extinction, de la structure dépendante de la fluorescence à auto-extinction, et une excellente efficacité photothermique. Dans notre formulation, porphysomes ont été synthétisés en utilisant l&#39;extrusion à haute pression, et présentaient une taille moyenne de particules d&#39;environ 120 nm. Injection de vingt-quatre heures après la intraveineuse de porphysomes, la température locale de la tumeur a augmenté de 30 ° C à 62 ° C rapidement lors d&#39;une exposition d&#39;une minute de 750 mW (1,18 W / cm 2), 671 nm irradiation laser. Suite à l&#39;ablation thermique complète de la tumeur, escarres formé et guéri en 2 semaines, alors que dans les groupes de contrôle les tumeurs ont continué à croître et à tous atteint le défini enpoint de d dans les 3 semaines. Ces données montrent comment porphysomes peuvent être utilisés comme thérapie photothermique puissant (PTT) des agents.

Pyropheophorbide est conjugué au phospholipide en tant que monomère de lipide stable (figure 1a) et la conjugués s&#39;auto-assembler pour former porphysomes par la méthode d&#39;extrusion de la membrane en utilisant une extrudeuse haute pression. Il est généralement difficile à extruder suspension porphysome-lipides pendant les trois premiers cycles de l&#39;extrusion à débit relativement lent. Comme de plus les extrusions sont répétées, le taux de solution extrudée d&#39;écoulement augme...

Watch this Scientific Journal Video about One Minute, Sub-One-Watt Photothermal Tumor Ablation Using Porphysomes, Intrinsic Multifunctional Nanovesicles at JoVE.com

One Minute, Sub-One-Watt Photothermal Tumor Ablation Using Porphysomes, Intrinsic Multifunctional Nanovesicles

Nous avons développé de nouveaux nanovésicules multifonctionnels appelés porphysomes intrinsèques, qui ont une structure dépendante fluorescence à auto-extinction et de propriétés uniques photothermiques, fonctionnant ainsi comme de puissants agents de thérapie photothermique. Nous avons formulé porphysomes utilisant extrusion à haute pression et étudié leur efficacité de la thérapie photothermique dans un modèle de xénogreffe de tumeur.

Une minute, Photothermal Tumor Ablation-un-Watt Sous Utilisation porphysomes, intrinsèque multifonctions nanovésicules

We recently developed porphysomes as intrinsically multifunctional nanovesicles. A photosensitizer, pyropheophorbide &alpha;, was conjugated to a ...

one-minute-sub-one-watt-photothermal-tumor-ablation-using-porphysomes

Research

JoVE Journal

Bioengineering

10.7K vues.  University of Toronto.  Nous avons développé de nouveaux nanovésicules multifonctionnels appelés porphysomes intrinsèques, qui ont une structure dépendante fluorescence à auto-extinction et de propriétés uniques photothermiques, fonctionnant ainsi comme de puissants agents de thérapie photothermique. Nous avons formulé porphysomes utilisant extrusion à haute pression et étudié leur efficacité de la thérapie photothermique dans un modèle de xénogreffe de tumeur. 

Vidéo: One Minute, Sub-One-Watt Photothermal Tumor Ablation Using Porphysomes, Intrinsic Multifunctional Nanovesicles

Contrast Ultrasound Targeted Treatment of Gliomas in Mice via Drug-Bearing Nanoparticle Delivery and Microvascular Ablation

We are developing minimally-invasive contrast agent microbubble based therapeutic approaches in which the permeabilization and/or ablation of the microvasculature are controlled by varying ultrasound pulsing parameters.  Specifically, we are testing whether such approaches may be used to treat malignant brain tumors through drug delivery and microvascular ablation.  Preliminary studies have been performed to determine whether targeted drug-bearing nanoparticle delivery can be facilitated by the ultrasound mediated destruction of "composite" delivery agents comprised of 100nm poly(lactide-co-glycolide) (PLAGA) nanoparticles that are adhered to albumin shelled microbubbles. We denote these agents as microbubble-nanoparticle composite agents (MNCAs). When targeted to subcutaneous C6 gliomas with ultrasound, we observed an immediate 4.6-fold increase in nanoparticle delivery in MNCA treated tumors over tumors treated with microbubbles co-administered with nanoparticles and a 8.5 fold increase over non-treated tumors. Furthermore, in many cancer applications, we believe it may be desirable to perform targeted drug delivery in conjunction with ablation of the tumor microcirculation, which will lead to tumor hypoxia and apoptosis. To this end, we have tested the efficacy of non-theramal cavitation-induced microvascular ablation, showing that this approach elicits tumor perfusion reduction, apoptosis, significant growth inhibition, and necrosis. Taken together, these results indicate that our ultrasound-targeted approach has the potential to increase therapeutic efficiency by creating tumor necrosis through microvascular ablation and/or simultaneously enhancing the drug payload in gliomas.

Insonation of microbubbles is a promising strategy for tumor ablation at reduced time-averaged acoustic powers, as well as for the targeted delivery of therapeutics. The purpose of the present study is to develop low duty cycle ultrasound pulsing strategies and nanocarriers to maximize non-thermal microvascular ablation and payload delivery to subcutaneous C6 gliomas.

L&#39;échographie de contraste ciblées traitement des gliomes chez la souris via la livraison de drogue nanoparticules-Bearing et l&#39;ablation microvasculaire

We are developing minimally-invasive contrast agent microbubble based therapeutic approaches in which the permeabilization and/or ablation of the ...

Recherche

Bio-ingénierie

Optical Frequency Domain Imaging of Ex vivo Pulmonary Resection Specimens: Obtaining One to One Image to Histopathology Correlation

Lung cancer is the leading cause of cancer-related deaths1. Squamous cell and small cell cancers typically arise in association with the conducting airways, whereas adenocarcinomas are typically more peripheral in location. Lung malignancy detection early in the disease process may be difficult due to several limitations: radiological resolution, bronchoscopic limitations in evaluating tissue underlying the airway mucosa and identifying early pathologic changes, and small sample size and/or incomplete sampling in histology biopsies. High resolution imaging modalities, such as optical frequency domain imaging (OFDI), provide non-destructive, large area 3-dimensional views of tissue microstructure to depths approaching 2 mm in real time (Figure 1)2-6. OFDI has been utilized in a variety of applications, including evaluation of coronary artery atherosclerosis6,7 and esophageal intestinal metaplasia and dysplasia6,8-10.
Bronchoscopic OCT/OFDI has been demonstrated as a safe in vivo imaging tool for evaluating the pulmonary airways11-23 (Animation). OCT has been assessed in pulmonary airways16,23 and parenchyma17,22 of animal models and in vivo human airway14,15. OCT imaging of normal airway has demonstrated visualization of airway layering and alveolar attachments, and evaluation of dysplastic lesions has been found useful in distinguishing grades of dysplasia in the bronchial mucosa11,12,20,21. OFDI imaging of bronchial mucosa has been demonstrated in a short bronchial segment (0.8 cm)18. Additionally, volumetric OFDI spanning multiple airway generations in swine and human pulmonary airways in vivo has been described19. Endobronchial OCT/OFDI is typically performed using thin, flexible catheters, which are compatible with standard bronchoscopic access ports. Additionally, OCT and OFDI needle-based probes have recently been developed, which may be used to image regions of the lung beyond the airway wall or pleural surface17.
While OCT/OFDI has been utilized and demonstrated as feasible for in vivo pulmonary imaging, no studies with precisely matched one-to-one OFDI:histology have been performed. Therefore, specific imaging criteria for various pulmonary pathologies have yet to be developed. Histopathological counterparts obtained in vivo consist of only small biopsy fragments, which are difficult to correlate with large OFDI datasets. Additionally, they do not provide the comprehensive histology needed for registration with large volume OFDI. As a result, specific imaging features of pulmonary pathology cannot be developed in the in vivo setting. Precisely matched, one-to-one OFDI and histology correlation is vital to accurately evaluate features seen in OFDI against histology as a gold standard in order to derive specific image interpretation criteria for pulmonary neoplasms and other pulmonary pathologies. Once specific imaging criteria have been developed and validated ex vivo with matched one-to-one histology, the criteria may then be applied to in vivo imaging studies. Here, we present a method for precise, one to one correlation between high resolution optical imaging and histology in ex vivo lung resection specimens. Throughout this manuscript, we describe the techniques used to match OFDI images to histology. However, this method is not specific to OFDI and can be used to obtain histology-registered images for any optical imaging technique. We performed airway centered OFDI with a specialized custom built bronchoscopic 2.4 French (0.8 mm diameter) catheter. Tissue samples were marked with tissue dye, visible in both OFDI and histology. Careful orientation procedures were used to precisely correlate imaging and histological sampling locations. The techniques outlined in this manuscript were used to conduct the first demonstration of volumetric OFDI with precise correlation to tissue-based diagnosis for evaluating pulmonary pathology24. This straightforward, effective technique may be extended to other tissue types to provide precise imaging to histology correlation needed to determine fine imaging features of both normal and diseased tissues.

Optical Frequency Domain Imaging of Ex vivo Pulmonary Resection Specimens: Obtaining One to One Image to Histopathology Correlation

A method to image ex vivo pulmonary resection specimens with optical frequency domain imaging (OFDI) and obtain precise correlation to histology is described, which is essential to developing specific OFDI interpretation criteria for pulmonary pathology. This method is applicable to other tissue types and imaging techniques to obtain precise imaging to histology correlation for accurate image interpretation and assessment. Imaging criteria established with this technique would then be applicable to image assessment in future in vivo studies.

D&#39;imagerie optique du domaine de fréquence<em&gt; Ex vivo</em&gt; Les spécimens résection pulmonaire: Obtention One to One image à l&#39;histopathologie Corrélation

Lung cancer is the leading cause of cancer-related deaths1. Squamous cell and small cell cancers typically arise in association with the conducting ...

Viral Nanoparticles for In vivo Tumor Imaging

The use of nanomaterials has the potential to revolutionize materials science and medicine. Currently, a number of different nanoparticles are being investigated for applications in imaging and therapy. Viral nanoparticles (VNPs) derived from plants can be regarded as self-assembled bionanomaterials with defined sizes and shapes. Plant viruses under investigation in the Steinmetz lab include icosahedral particles formed by Cowpea mosaic virus (CPMV) and Brome mosaic virus (BMV), both of which are 30 nm in diameter. We are also developing rod-shaped and filamentous structures derived from the following plant viruses: Tobacco mosaic virus (TMV), which forms rigid rods with dimensions of 300 nm by 18 nm, and Potato virus X (PVX), which form filamentous particles 515 nm in length and 13 nm in width (the reader is referred to refs. 1 and 2 for further information on VNPs).
From a materials scientist's point of view, VNPs are attractive building blocks for several reasons: the particles are monodisperse, can be produced with ease on large scale in planta, are exceptionally stable, and biocompatible. Also, VNPs are "programmable" units, which can be specifically engineered using genetic modification or chemical bioconjugation methods 3. The structure of VNPs is known to atomic resolution, and modifications can be carried out with spatial precision at the atomic level4, a level of control that cannot be achieved using synthetic nanomaterials with current state-of-the-art technologies.
In this paper, we describe the propagation of CPMV, PVX, TMV, and BMV in Vigna ungiuculata and Nicotiana benthamiana plants. Extraction and purification protocols for each VNP are given. Methods for characterization of purified and chemically-labeled VNPs are described. In this study, we focus on chemical labeling of VNPs with fluorophores (e.g. Alexa Fluor 647) and polyethylene glycol (PEG). The dyes facilitate tracking and detection of the VNPs 5-10, and PEG reduces immunogenicity of the proteinaceous nanoparticles while enhancing their pharmacokinetics 8,11. We demonstrate tumor homing of PEGylated VNPs using a mouse xenograft tumor model. A combination of fluorescence imaging of tissues ex vivo using Maestro Imaging System, fluorescence quantification in homogenized tissues, and confocal microscopy is used to study biodistribution. VNPs are cleared via the reticuloendothelial system (RES); tumor homing is achieved passively via the enhanced permeability and retention (EPR) effect12. The VNP nanotechnology is a powerful plug-and-play technology to image and treat sites of disease in vivo. We are further developing VNPs to carry drug cargos and clinically-relevant imaging moieties, as well as tissue-specific ligands to target molecular receptors overexpressed in cancer and cardiovascular disease.

Viral Nanoparticles for In vivo Tumor Imaging

Plant viral nanoparticles (VNPs) are promising platforms for applications in biomedicine. Here, we describe the procedures for plant VNP propagation, purification, characterization, and bioconjugation. Finally, we show the application of VNPs for tumor homing and imaging using a mouse xenograft model and fluorescence imaging.

Les nanoparticules virales pour<em&gt; In vivo</em&gt; Imagerie des tumeurs

The use of nanomaterials has the potential to revolutionize materials science and medicine. Currently, a number of different nanoparticles are being ...

Construction of a Preclinical Multimodality Phantom Using Tissue-mimicking Materials for Quality Assurance in Tumor Size Measurement

World Health Organization (WHO) and the Response Evaluation Criteria in Solid Tumors (RECIST) working groups advocated standardized criteria for radiologic assessment of solid tumors in response to anti-tumor drug therapy in the 1980s and 1990s, respectively. WHO criteria measure solid tumors in two-dimensions, whereas RECIST measurements use only one-dimension which is considered to be more reproducible 1, 2, 3,4,5. These criteria have been widely used as the only imaging biomarker approved by the United States Food and Drug Administration (FDA) 6. In order to measure tumor response to anti-tumor drugs on images with accuracy, therefore, a robust quality assurance (QA) procedures and corresponding QA phantom are needed.
To address this need, the authors constructed a preclinical multimodality (for ultrasound (US), computed tomography (CT) and magnetic resonance imaging (MRI)) phantom using tissue-mimicking (TM) materials based on the limited number of target lesions required by RECIST by revising a Gammex US commercial phantom 7. The Appendix in Lee et al. demonstrates the procedures of phantom fabrication 7. In this article, all protocols are introduced in a step-by-step fashion beginning with procedures for preparing the silicone molds for casting tumor-simulating test objects in the phantom, followed by preparation of TM materials for multimodality imaging, and finally construction of the preclinical multimodality QA phantom. The primary purpose of this paper is to provide the protocols to allow anyone interested in independently constructing a phantom for their own projects. QA procedures for tumor size measurement, and RECIST, WHO and volume measurement results of test objects made at multiple institutions using this QA phantom are shown in detail in Lee et al. 8.

This paper describes in-house procedures of constructing a preclinical multimodality phantom made of tissue-mimicking (TM) materials for quality assurance (QA) of tumor size measurement in animal imaging modalities such as ultrasound (US), computed tomography (CT) and magnetic resonance imaging (MRI).

Construction d&#39;un fantôme de multimodalité préclinique employant des matériaux tissus imitant pour l&#39;assurance qualité dans la mesure de la taille des tumeurs

World Health Organization (WHO) and the Response Evaluation Criteria in Solid Tumors (RECIST) working groups advocated standardized criteria for ...

Production of Large Numbers of Size-controlled Tumor Spheroids Using Microwell Plates

Tumor spheroids are increasingly recognized as an important in vitro model for the behavior of tumor cells in three dimensions. More physiologically relevant than conventional adherent-sheet cultures, they more accurately recapitulate the complexity and interactions present in real tumors. In order to harness this model to better assess tumor biology, or the efficacy of novel therapeutic agents, it is necessary to be able to generate spheroids reproducibly, in a controlled manner and in significant numbers.
The AggreWell system consists of a high-density array of pyramid-shaped microwells, into which a suspension of single cells is centrifuged. The numbers of cells clustering at the bottom of each microwell, and the number and ratio of distinct cell types involved depend only on the properties of the suspension introduced by the experimenter. Thus, we are able to generate tumor spheroids of arbitrary size and composition without needing to modify the underlying platform technology. The hundreds of microwells per square centimeter of culture surface area in turn ensure that extremely high production levels may be attained via a straightforward, nonlabor-intensive process. We therefore expect that this protocol will be broadly useful to researchers in the tumor spheroid field.

We introduce a protocol for the generation of large numbers (thousands to hundreds of thousands) of uniform size- and composition-controlled tumor spheroids, using commercially available microwell plates.

La production de grands nombres de tumeurs Sphéroïdes Taille contrôlée à l&#39;aide de micro puits Plaques

Tumor spheroids are increasingly recognized as an important in vitro model for the behavior of tumor cells in three dimensions. More physiologically ...

PLGA Nanoparticles Formed by Single- or Double-emulsion with Vitamin E-TPGS

Poly(lactic-co-glycolic acid) (PLGA) is a biocompatible member of the aliphatic polyester family of biodegradable polymers. PLGA has long been a popular choice for drug delivery applications, particularly since it is already FDA-approved for use in humans in the form of resorbable sutures. Hydrophobic and hydrophilic drugs are encapsulated in PLGA particles via single- or double-emulsion. Briefly, the drug is dissolved with polymer or emulsified with polymer in an organic phase that is then emulsified with the aqueous phase. After the solvent has evaporated, particles are washed and collected via centrifugation for lyophilization and long term storage. PLGA degrades slowly via hydrolysis in aqueous environments, and encapsulated agents are released over a period of weeks to months. Although PLGA is a material that possesses many advantages for drug delivery, reproducible formation of nanoparticles can be challenging; considerable variability is introduced by the use of different equipment, reagents batch, and precise method of emulsification. Here, we describe in great detail the formation and characterization of microparticles and nanoparticles formed by single- or double-emulsion using the emulsifying agent vitamin E-TPGS. Particle morphology and size are determined with scanning electron microscopy (SEM). We provide representative SEM images for nanoparticles produced with varying emulsifier concentration, as well as examples of imaging artifacts and failed emulsifications. This protocol can be readily adapted to use alternative emulsifiers (e.g. poly(vinyl alcohol), PVA) or solvents (e.g.&#160;dichloromethane, DCM).

We describe the production and characterization of nanoparticles and microparticles composed of poly(lactic-co-glycolic acid) using vitamin E-TPGS as an emulsifier. By varying formulation parameters such as the concentration of emulsifier, it is possible to produce nanoparticles with mean diameters ranging from 220 nm to&#160;1.98 &#181;m.

Poly(lactic-co-glycolic acid) (PLGA) is a biocompatible member of the aliphatic polyester family of biodegradable polymers. PLGA has long been a popular ...

Longitudinal Measurement of Extracellular Matrix Rigidity in 3D Tumor Models Using Particle-tracking Microrheology

The mechanical microenvironment has been shown to act as a crucial regulator of tumor growth behavior and signaling, which is itself remodeled and modified as part of a set of complex, two-way mechanosensitive interactions. While the development of biologically-relevant 3D tumor models have facilitated mechanistic studies on the impact of matrix rheology on tumor growth, the inverse problem of mapping changes in the mechanical environment induced by tumors remains challenging. Here, we describe the implementation of particle-tracking microrheology (PTM) in conjunction with 3D models of pancreatic cancer as part of a robust and viable approach for longitudinally monitoring physical changes in the tumor microenvironment, in situ. The methodology described here integrates a system of preparing in vitro 3D models embedded in a model extracellular matrix (ECM) scaffold of Type I collagen with fluorescently labeled probes uniformly distributed for position- and time-dependent microrheology measurements throughout the specimen. In vitro tumors are plated and probed in parallel conditions using multiwell imaging plates. Drawing on established methods, videos of tracer probe movements are transformed via the Generalized Stokes Einstein Relation (GSER) to report the complex frequency-dependent viscoelastic shear modulus, G*(&#969;). Because this approach is imaging-based, mechanical characterization is also mapped onto large transmitted-light spatial fields to simultaneously report qualitative changes in 3D tumor size and phenotype. Representative results showing contrasting mechanical response in sub-regions associated with localized invasion-induced matrix degradation as well as system calibration, validation data are presented. Undesirable outcomes from common experimental errors and troubleshooting of these issues are also presented. The 96-well 3D culture plating format implemented in this protocol is conducive to correlation of microrheology measurements with therapeutic screening assays or molecular imaging to gain new insights into impact of treatments or biochemical stimuli on the mechanical microenvironment.

Particle-tracking microrheology can be used to non-destructively quantify and spatially map changes in extracellular matrix mechanical properties in 3D tumor models.

Mesure longitudinale de la matrice extracellulaire rigidité de tumeur modèles 3D utilisant Microrhéologie de particules de suivi

The mechanical microenvironment has been shown to act as a crucial regulator of tumor growth behavior and signaling, which is itself remodeled and ...

Real-time Monitoring of High Intensity Focused Ultrasound (HIFU) Ablation of In Vitro Canine Livers Using Harmonic Motion Imaging for Focused Ultrasound (HMIFU)

Harmonic Motion Imaging for Focused Ultrasound (HMIFU) is a technique that can perform and monitor high-intensity focused ultrasound (HIFU) ablation. An oscillatory motion is generated at the focus of a 93-element and 4.5 MHz center frequency HIFU transducer by applying a 25 Hz amplitude-modulated signal using a function generator. A 64-element and 2.5 MHz imaging transducer with 68kPa peak pressure is confocally placed at the center of the HIFU transducer to acquire the radio-frequency (RF) channel data. In this protocol, real-time monitoring of thermal ablation using HIFU with an acoustic power of 7 W on canine livers in vitro is described. HIFU treatment is applied on the tissue during 2 min and the ablated region is imaged in real-time using diverging or plane wave imaging up to 1,000 frames/second. The matrix of RF channel data is multiplied by a sparse matrix for image reconstruction. The reconstructed field of view is of 90&#176; for diverging wave and 20 mm for plane wave imaging and the data are sampled at 80 MHz. The reconstruction is performed on a Graphical Processing Unit (GPU) in order to image in real-time at a 4.5 display frame rate. 1-D normalized cross-correlation of the reconstructed RF data is used to estimate axial displacements in the focal region. The magnitude of the peak-to-peak displacement at the focal depth decreases during the thermal ablation which denotes stiffening of the tissue due to the formation of a lesion. The displacement signal-to-noise ratio (SNRd) at the focal area for plane wave was 1.4 times higher than for diverging wave showing that plane wave imaging appears to produce better displacement maps quality for HMIFU than diverging wave imaging.

Real-time Monitoring of High Intensity Focused Ultrasound HIFU Ablation of In Vitro Canine Livers Using Harmonic Motion Imaging for Focused Ultrasound HMIFU

This article describes real-time monitoring of HIFU ablation in canine liver with high frame rate ultrasound imaging using diverging and plane wave imaging. Harmonic Motion Imaging for Focused Ultrasound is used to image the decrease of acoustic radiation force induced displacement in the ablated region.

Suivi en temps réel de haute intensité ultrasons focalisés (HIFU) Ablation de<em&gt; In Vitro</em&gt; Canine Livers Utilisation mouvement harmonique Imaging pour ultrasons focalisés (HMIFU)

Harmonic Motion Imaging for Focused Ultrasound (HMIFU) is a technique that can perform and monitor high-intensity focused ultrasound (HIFU) ablation. An ...

Tissue Engineering by Intrinsic Vascularization in an In Vivo Tissue Engineering Chamber

In reconstructive surgery, there is a clinical need for an alternative to the current methods of autologous reconstruction which are complex, costly and trade one defect for another. Tissue engineering holds the promise to address this increasing demand. However, most tissue engineering strategies fail to generate stable and functional tissue substitutes because of poor vascularization. This paper focuses on an in vivo tissue engineering chamber model of intrinsic vascularization where a perfused artery and a vein either as an arteriovenous loop or a flow-through pedicle configuration is directed inside a protected hollow chamber. In this chamber-based system angiogenic sprouting occurs from the arteriovenous vessels and this system attracts ischemic and inflammatory driven endogenous cell migration which gradually fills the chamber space with fibro-vascular tissue. Exogenous cell/matrix implantation at the time of chamber construction enhances cell survival and determines specificity of the engineered tissues which develop. Our studies have shown that this chamber model can successfully generate different tissues such as fat, cardiac muscle, liver and others. However, modifications and refinements are required to ensure target tissue formation is consistent and reproducible. This article describes a standardized protocol for the fabrication of two different vascularized tissue engineering chamber models in vivo.

Tissue Engineering by Intrinsic Vascularization in an In Vivo Tissue Engineering Chamber

This is a guideline for constructing in vivo vascularized tissue using a microsurgical arteriovenous loop or a flow-through pedicle configuration inside a tissue engineering chamber. The vascularized tissues generated can be employed for organ regeneration and replacement of tissue defects, as well as for drug testing and disease modeling.

Génie tissulaire par Intrinsic vascularisation dans un<em&gt; In Vivo</em&gt; Tissue Chambre Ingénierie

In reconstructive surgery, there is a clinical need for an alternative to the current methods of autologous reconstruction which are complex, costly and ...

A Rapid and Chemical-free Hemoglobin Assay with Photothermal Angular Light Scattering

Photo-thermal angular light scattering (PT-AS) is a novel optical method for measuring the hemoglobin concentration ([Hb]) of blood samples. On the basis of the intrinsic photothermal response of hemoglobin molecules, the sensor enables high-sensitivity, chemical-free measurement of [Hb]. [Hb] detection capability with a limit of 0.12 g/dl over the range of 0.35 - 17.9 g/dl has been demonstrated previously. The method can be readily implemented using inexpensive consumer electronic devices such as a laser pointer and a webcam. The use of a micro-capillary tube as a blood container also enables the hemoglobin assay with a nanoliter-scale blood volume and a low operating cost. Here, detailed instructions for the PT-AS optical setup and signal processing procedures are presented. Experimental protocols and representative results for blood samples in anemic conditions ([Hb] = 5.3, 7.5, and 9.9 g/dl) are also provided, and the measurements are compared with those from a hematology analyzer. Its simplicity in implementation and operation should enable its wide adoption in clinical laboratories and resource-limited settings.

A photo-thermal angular light scattering (PT-AS) sensor enables the rapid and chemical-free hemoglobin assay of nanoliter-scale blood samples. Here, details of the PT-AS setup and a measurement protocol for the hemoglobin concentration in blood are provided. Representative results for anemic blood samples are also presented.

A Hémoglobine Assay rapide et sans produits chimiques avec Photothermique angulaire Light Scattering

Photo-thermal angular light scattering (PT-AS) is a novel optical method for measuring the hemoglobin concentration ([Hb]) of blood samples. On the basis ...