Yazarlar yazının Dr. Lynnea Brumbaugh yakın okuma için teşekkür ederiz. Bu çalışma Ulusal Sağlık Grants R01 EB000712, R01 EB008085, R01 CA134539, U54 CA136398 ve 5P60 DK02057933 Enstitüleri tarafından sponsor oldu. Prof Lihong V. Wang Microphotoacoustics, Inc ve Endra, Inc Ancak, bu işe destek vermedi, bir finansal çıkarı vardır.

1. Sistem yapılandırması <ol><li> Optik ışınlama <ol><li> Optik ışınlama kaynağı: solid-state darbeli lazer (INNOSLAB, Edgewave) ve dye lazer (CBR-Ge, Sirah) pompalanır-diyot. </li><li> Çıkış lazer ışını (darbe genişliği: 7 ns) (p50c, Thorlabs) 50 mikron iğne deliğinden geçirmek için yoğunlaştırıcı mercek (LA1131, Thorlabs) odaklanmıştır. </li><li> Iğne deliği iğne deliği çapı etkin bir mekansal filtreleme için temel mod ışın çapı ile maç kondenser lens odak biraz uzakta konumlandırılmış. </li><li> Süzülmüş ışını bir tek modlu optik fiber (P1-460A-FC-2, Thorlabs) içine birleştiğinde sonra bir nötr yoğunluk filtresi (NDC-50C-2M, Thorlabs) ve zayıflatılmış. </li><li> 570 nm dalga boyunda bir kırınım sınırlı ~ 2.6 mm optik odak fiber çıkış elde etmek için bir mikroskop objektif arka diyaframı (RMS4X, Thorlabs) doldurur. </li></ol></li><li> Ultrasonik algılama <ol><li> Ultrasonik transdüser: 50-MHz merkez frekansı (V214-BB-RM, Olympus-NDT). </li><li> Ultrasonik sensör, ultrasonik algılama, kırınım sınırlı optik radyoterapi ile aynı eksende hizalanmış bir ev yapımı akustik-optik ışın birleştirici 11 eklenir . </li><li> Küresel kavite akustik lens üretmek için birleştirici alt zemin. Bu akustik lens, su, 0.5 &#39;lik bir sayısal diyafram ve 50-MHz merkez frekansı akustik bir odak çapı 43 mikron verir. </li><li> Optik ve akustik odaklar algılama duyarlılığını en üst düzeye çıkarmak için confocally hizalanır. </li></ol></li><li> Akustik kaplin <ol><li> Kuru ultrasonik kaplin erken foto akustik görüntüleme sistemleri 12 olduğu, su, deney hayvanlarında daldırarak önlemek için kullanılmaktadır . </li><li> Petri kabı (çapı 9 cm) altında bir görüntüleme penceresi açılır ve bir ultrasonik ve optik olarak saydam polietilen membran ile mühürlenir. </li><li> Polietilen membran ve görüntülü çiftler batık OR-PAM görüntüleme baş Petri kabındaki Petri kabı nesneden oluşturulan foto akustik dalga ve deiyonize su dalgası daha fazla çiftler nesne arasındaki Ultrasonik jel (Temiz görüntü SonoTech) . </li></ol></li><li> Elektronik <ol><li> Ultrasonik sensör tarafından tespit foto akustik sinyal iki basamaklanmasını yükselticiler (ZFL 500LN, Mini-Devreleri) tarafından yükseltilir. </li><li> Güçlendirilmiş sinyal 200 MS / s örnekleme oranı 14-bit veri toplama (DAQ) kartı (14200 CompuScope Gage Uygulamalı Bilimler) dijitalleştirilmiştir </li></ol></li><li> Şeması Tarama <ol><li> Raster, yatay (xy) düzlemi boyunca OR-PAM görüntüleme kafa tarama İki boyutlu (2-D) DAQ kurulu ve pompa lazer hem de tetikleyen bir kişisel bilgisayar tarafından kontrol edilir. Tetikleme sinyali DAQ kurulu saat sinyali ile senkronize edilir. </li><li> 2-B tarayıcının hızlı eksen kesitsel tarama (B-scan) yönü olarak tanımlanır. </li><li> B-tarama görüntü dizisi, doğrudan 3 boyutlu render veya 2-D maksimum genlik projeksiyon (MAP) görüntü ya görülebilir hacimsel bir görüntü oluşturmak için yavaş eksen boyunca görüntüleme kafasını çevirerek elde edilebilir . </li></ol></li></ol> 2. Sistem hizalama <ol><li> Akustik odak düzlemli konumunu belirlemek için (yani, maksimum darbe-yankı ultrasonik sinyali tetikleme sinyali zaman gecikmesi) darbe-eko ultrason ve ultrasonik reflektör kullanın. Bu adım, OR-PAM sistemi oluştururken sadece bir kez yapılması gereklidir. </li><li> Single-mode fiber bağlantı verimliliği maksimize. </li><li> Optik emici bir nesne (örneğin, bir parça siyah bant) üst ultrasonik jel uygulayın ve hafifçe deiyonize su ile doldurulur Petri kabındaki görüntüleme penceresi altına ekleyin. </li><li> Görüntüleme kafası suya indirin ve akustik mercek altında hava kabarcığı kaldırmak. </li><li> Emici nesne foto akustik sinyal görüntüleme kafası akustik gecikme değerlendirilecektir akustik odak düzlemli kadar ayarlayın. </li><li> Düz nesneden oluşturulan foto akustik sinyal genliği en üst düzeye çıkarmak için mikroskop objektif dikey konumda (yani, z konum) ayarlayın. Maksimize sinyal genliği, optik odak dikey yönde akustik odak ile uyumlu olduğunu göstermektedir. </li><li> Mikroskop amacı yatay pozisyonları (yani, x ve y pozisyonlarını) hedef üretilen foto akustik sinyal simetrik desen görünene kadar ayarlayın. Simetri optik odak yatay yönde akustik odak ile uyumlu olduğunu göstermektedir. </li><li> Foto akustik sinyal simetri ve genlik hem de optimize kadar 2.6 ve 2.7 adımları tekrarlayın. </li></ol> 3. A sa<html> mple deneysel işlemin in vivo OR-PAM fare kulak vasküler <ol><li> Bu adım, çıplak fareler için gerekli değildir . Bir kokteyl intraperitoneal [:: 0.1 ml/10 g dozu 1 ml ketamin (100 mg / ml), 0.1 ml ksilizin (100 mg / ml) ve 8.9 ml serum fizyolojik tarifi] hayvan anestezisi. Kulak saçlarını tıraş ve daha fazla cerrahi Krem (Kategori: 82.565, American International Industries) ile kalan saç kıllarını deiyonize su ile temizlemeden önce. Gibi epilasyon deri vaskülatürü hafifçe tahriş olabilir ve böylece en iyi 24 saat önce planlanan deney yapılır. </li><li> Foto akustik lazer sistemi açın ve sistem hizalamayı kontrol edin. </li><li> Izofluran% 3 buharlaşmış inhalasyon gaz (tipik akış hızı, hayvanın vücut ağırlığına bağlı olarak, 1.0-1.5 lt / dk) ile fare anestezi ve deney boyunca% 1 izofluran ile anestezi korumak. Medikal dereceli hava fare normal fizyolojik durumunu korumak için inhalasyon gaz olarak tavsiye edilir. </li><li> Stereotaktik bir aşamaya fare aktarın ve bir ısıtma yastığı ile vücut ısısı 37 ° ° C kontrol. </li><li> Plastik bir plaka üzerinde Düzleştir fare kulak ve kulak üstündeki ultrason jel tabakası uygulayın. Jel içindeki hava kabarcıklarını yakalama kaçının. Daha sonra, görüntüleme penceresi altında kulağı ve ultrason jel temas polietilen membran alt kadar yavaş yavaş hayvan aşamada yükseltmek. Yumuşak kontakt karşı kulak zarı basarak kulağa kan akışını etkileyebilir, çünkü gereklidir. </li><li> Fizyolojik durumunu izlemek için fare bacak veya kuyruk bir pulse oksimetre Kelepçe, kuruluk ve yanlışlıkla lazer fare gözleri hasarı önlemek için gözlere merhem uygulayın. </li><li> Akustik lens deiyonize suya batırılır kadar görüntüleme kafasını indirin ve akustik mercek altında hava kabarcığı kaldırmak. </li><li> Lazer fluens Amerikan Ulusal Standartlar Enstitüsü 13 lazer güvenlik standartları içinde olduğundan emin olmak için kontrol edin. Lazer akıcılık, lazer ışını, cilt yüzeyi altındaki 150 mm odaklı lazer darbe enerji 80 NJ çeviren 20 mJ / cm 2, geçmemelidir. </li><li> Lazer harici tetikleme modu ayarlama ve deneme taramayı başlatabilirsiniz. Güçlü foto akustik sinyal akustik odak düzlemli kadar görüntüleme kafa z konumunu ayarlayın. </li><li> Doğru tarama parametrelerini ayarlayın ve resmi görüntü alımı başlatabilirsiniz. </li><li> Deney sonucunda, lazer kapatmak, deiyonize su görüntüleme kafasını dışarı çıkarın, fareyi bırakın hayvan evresini, fare kulağı deiyonize su ile temizlemek, anestezi sistemi ve sıcaklık kontrolörü kapatın ve fare boşaltma stereotaktik sahneden. </li><li> Tekrarlayan görüntüleme gerekli ise, 37 olarak belirlenmiştir çevre sıcaklığı ° C ile bir kuluçka fare koymak Doğal olarak uyanır sonra fare hayvan tesis dönün. Aksi takdirde, euthanatize ve bertaraf hayvan protokolleri uygulayın. </li></ol> 4. Fonksiyonel OR-PAM toplam konsantrasyonu ve hemoglobin oksijen satürasyonu <ol><li> Oksihemoglobin (HBO 2) ve deoksihemoglobin (HBR), hemoglobin iki büyük formlar, görünür spektral aralıkta baskın endojen foto akustik kaynak vardır . HBO 2 ve HBR farklı optik soğurma spektrumları ve böylece toplam konsantrasyonu (HBT) ve 5 hemoglobin oksijen doygunluğu (SO 2) Işıksal ölçmek için ayırt edilebilir. İşte sO 2 ölçüm için uygun optik dalga boylarında seçmenize yardımcı olmak için iki kurallar şunlardır : </li><li> Q-bant içinde hemoglobin absorbsiyon spektrumunun (yani, 550-600 nm) dalga boylarında yeterli bir sinyal-gürültü oranı ve yeterli penetrasyon sağlamak için seçilmiş olmalıdır. </li><li> HBO 2 ve HBR emme katsayıları belirgin bir fark vardır (örneğin, HBR-egemen 561 nm ve HBO 2-baskın 578 nm) dalga boylarında tavsiye edilir. </li></ol> SO 2 yanı sıra, HBT ekleyerek hesaplanabilir [HBR] ve [HBO 2] birlikte veya doğrudan hemoglobin molar sönüm katsayısı spektrumları (örneğin, 530 nm, 545 nm, 570 nm ve isosbestic dalga boylarında ölçülen olabilir HBR ve HBO 2 eşit molar yok katsayıları 584 nm) 14,. 5. Temsilcisi Sonuçlar: Şekil 1&#39;de gösterildiği gibi çift dalga boyu (561 ve 570 nm) OR-PAM tarafından görüntülü canlı çıplak bir fare kulak vasküler anatomi ve 2 sO. ~ 80 dakika:: ilgi, böyle bir bölgede 2 ölçümleri (2.5 mm x 5 mm adım boyutu 10 mm x 10 mm görüntü boyutu) sO çift dalga boyu için tipik bir görüntü elde etme zamanı . p_upload/2729/2729fig1.jpg &quot;alt =&quot; Şekil 1 &quot;/&gt; Şekil 1 in vivo optik çözünürlük foto akustik mikroskopi. (A) (570 nm edinilen) vasküler anatomi gösteren toplam hemoglobin konsantrasyonu ve (B) çıplak bir fare kulak hemoglobin oksijen satürasyonu (561 nm ve 570 nm edinilmiş) HARİTASI görüntüler. (C) Close-up panelinde kutulu bölge (A). Panelindeki ölçekli bar (A) hem (A) geçerlidir ve (B).

<ol>
	<li>Maslov, K., Zhang, H. F., Hu, S., Wang, L. V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Optical-resolution+photoacoustic+microscopy+for+in+vivo+imaging+of+single+capillaries.">Optical-resolution photoacoustic microscopy for in vivo imaging of single capillaries.</a> Opt. Lett. 33, 929-931 (2008).</li><li>Zhang, C., Maslov, K., Wang, L. V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Subwavelength-resolution+label-free+photoacoustic+microscopy+of+optical+absorption+in+vivo.">Subwavelength-resolution label-free photoacoustic microscopy of optical absorption in vivo.</a> Opt. Lett. 35, 3195-3197 (2010).</li><li>Wang, Y., Maslov, K., Kim, C., Hu, S., Wang, L. V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Integrated+photoacoustic+and+fluorescence+confocal+microscopy.">Integrated photoacoustic and fluorescence confocal microscopy.</a> IEEE. Trans. Biomed. Eng. 57, 2576-2578 (2010).</li><li>Jiao, S., Xie, Z., Zhang, H. F., Puliafito, C. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Simultaneous+multimodal+imaging+with+integrated+photoacoustic+microscopy+and+optical+coherence+tomography.">Simultaneous multimodal imaging with integrated photoacoustic microscopy and optical coherence tomography.</a> Opt. Lett. 34, 2961-2963 (2009).</li><li>Hu, S., Maslov, K., Tsytsarev, V., Wang, L. 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Tüm deneysel hayvan prosedürleri School of St Louis Washington Üniversitesi Tıp Hayvan Araştırmaları Komitesi tarafından onaylanmış laboratuar hayvan protokolü ile uygun olarak gerçekleştirilmiştir.

Bu video, OR-PAM deneysel protokoller, sistem konfigürasyonuna, sistem hizalama ve tipik deneysel prosedürleri de dahil olmak üzere ayrıntılı bir talimat sağlar. Etiket içermeyen, non-invaziv OR-PAM tek bir kılcal bazında mikrovasküler işleyişi ve metabolizma çalışmaları etkin ve böylece mikrosirkülasyon ilgili fizyoloji ve patoloji anlayışımız genişletmek için bir potansiyele sahiptir. Microphotoacoustics OR-PAM sistemi şu anda bu üretim yapmaktadır.

Ev yapımı akustik-optik ışın birleştirici: <ul><li> dik açılı prizma (NT32-545, Edmund Optik) </li><li> rhomboid prizma (NT49-419, Edmund Optik) </li><li> silikon yağı (1000cSt, Clearco Ürünler) </li><li> OR-PAM sistemi (Microphotoacoustics) </li></ul>

three-dimensional optical-resolution photoacoustic microscopy

Organel ve hücresel düzeyde değerli bilgiler sağlayan optik mikroskop, çok yetenekli bir biyomedikal teknoloji olarak kabul edilmiştir. Olarak in vivo olarak üç boyutlu (3-D), optik mikroskop, single-/multi-photon floresan mikroskopi ve Optik Koherens Tomografi (OCT), sırasıyla olağanüstü duyarlılıkları, floresan ve optik saçılma zıtlıklar göstermiştir dayanaklarından. Ancak, temel fizyolojik / patolojik bilgileri kodlayan, biyolojik dokular, optik soğurma kontrastı henüz değerlendirilebilir edilmiş değil. Biyomedikal photoacoustics ortaya çıkışı, optik ışınlama hücresel 1 veya 2 düzeyinde yatay çözünürlük bile Subselüler ulaşmak için kırınım sınırı odaklı optik mikroskop optik çözünürlükte foto akustik mikroskobu (OR-PAM) 1, yeni bir şube açmıştır . OR-PAM değerli bir tamamlayıcısı olarak, mevcut optik mikroskop teknolojileri, en az iki yenilikler getiriyor. Birinci ve en önemlisi, OR-PAM olağanüstü hassasiyeti (yani% 100) ile optik soğurma kontrastlar algılar. Floresan mikroskobu 3 veya optik saçılma tabanlı Ekim 4 (veya her ikisi ile) ile OR-PAM birleştiren kapsamlı biyolojik dokuların optik özellikleri sağlar. İkincisi, floresan mikroskopi ve OCT saf optik süreçler aksine OR-PAM, akustik dalgalar, optik soğurma kodlar, arka plan ücretsiz algılanmasını sağlar. OR-PAM akustik algılama sinyal bozulması optik saçılma etkileri azaltır ve doğal olarak uyarma, uyarma ve floresans spektrumları arasındaki örtüşme nedeniyle floresan mikroskobu yaygın bir sorundur algılama arasındaki olası etkileşimler (yani, crosstalks) ortadan kaldırır. Benzersiz optik soğurma görüntüleme için, OR-PAM dahil olmak üzere, icadından bu yana geniş biyomedikal uygulamalar göstermiştir, fakat nöroloji 5, 6, 7 oftalmoloji, 8, vasküler biyoloji 9 ve 10 dermatoloji, sınırlı değildir. Bu video, sistem konfigürasyonu ve OR-PAM uyumun yanı sıra, in vivo fonksiyonel mikrovasküler görüntüleme deneysel prosedürleri öğretir .

Watch this Scientific Journal Video about Three-dimensional Optical-resolution Photoacoustic Microscopy at JoVE.com

Three-dimensional Optical-resolution Photoacoustic Microscopy

Optik çözünürlüklü foto akustik mikroskobu (OR-PAM), görüntüleme optik soğurma yeteneğine sahip gelişmekte olan bir teknoloji tezat<em&gt; In vivo</em&gt; Hücresel çözünürlük ve hassasiyet. İşte, biz, sistem konfigürasyonuna, sistem hizalama da dahil olmak üzere, OR-PAM deneysel protokolleri, görüntülenmiştir bir talimat tipik<em&gt; In vivo</em&gt; Deneysel prosedürleri ve fonksiyonel görüntüleme programları.

Üç boyutlu optik çözünürlük foto akustik Mikroskopi

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

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17.7K Görüntüleme.  Washington University in St. Louis. Optik çözünürlüklü foto akustik mikroskobu (OR-PAM), görüntüleme optik soğurma yeteneğine sahip gelişmekte olan bir teknoloji tezat In vivo Hücresel çözünürlük ve hassasiyet. İşte, biz, sistem konfigürasyonuna, sistem hizalama da dahil olmak üzere, OR-PAM deneysel protokolleri, görüntülenmiştir bir talimat tipik In vivo Deneysel prosedürleri ve fonksiyonel görüntüleme programları.

Video: Three-dimensional Optical-resolution Photoacoustic Microscopy

Alginate Hydrogels for Three-Dimensional Organ Culture of Ovaries and Oviducts

Ovarian cancer is the fifth leading cause of cancer deaths in women and has a 63% mortality rate in the United States1. The cell type of origin for ovarian cancers is still in question and might be either the ovarian surface epithelium (OSE) or the distal epithelium of the fallopian tube fimbriae2,3. Culturing the normal cells as a primary culture in vitro will enable scientists to model specific changes that might lead to ovarian cancer in the distinct epithelium, thereby definitively determining the cell type of origin. This will allow development of more accurate biomarkers, animal models with tissue-specific gene changes, and better prevention strategies targeted to this disease.
Maintaining normal cells in alginate hydrogels promotes short term in vitro culture of cells in their three-dimensional context and permits introduction of plasmid DNA, siRNA, and small molecules. By culturing organs in pieces that are derived from strategic cuts using a scalpel, several cultures from a single organ can be generated, increasing the number of experiments from a single animal. These cuts model aspects of ovulation leading to proliferation of the OSE, which is associated with ovarian cancer formation. Cell types such as the OSE that do not grow well on plastic surfaces can be cultured using this method and facilitate investigation into normal cellular processes or the earliest events in cancer formation4.
Alginate hydrogels can be used to support the growth of many types of tissues5. Alginate is a linear polysaccharide composed of repeating units of &beta;-D-mannuronic acid and &alpha;-L-guluronic acid that can be crosslinked with calcium ions, resulting in a gentle gelling action that does not damage tissues6,7. Like other three-dimensional cell culture matrices such as Matrigel, alginate provides mechanical support for tissues; however, proteins are not reactive with the alginate matrix, and therefore alginate functions as a synthetic extracellular matrix that does not initiate cell signaling5. The alginate hydrogel floats in standard cell culture medium and supports the architecture of the tissue growth in vitro.
A method is presented for the preparation, separation, and embedding of ovarian and oviductal organ pieces into alginate hydrogels, which can be maintained in culture for up to two weeks. The enzymatic release of cells for analysis of proteins and RNA samples from the organ culture is also described. Finally, the growth of primary cell types is possible without genetic immortalization from mice and permits investigators to use knockout and transgenic mice.

Culture of normal cells in their three-dimensional context represents an alternative method to study early events required for cellular transformation and tumorigenesis. This method is used to grow normal ovarian and oviductal cells to study early events in ovarian cancer formation.

Yumurtalıklar ve fallop borularında Üç Boyutlu Organ Kültür Aljinatta Hidrojeller

Ovarian cancer is the fifth leading cause of cancer deaths in women and has a 63% mortality rate in the United States1. The cell type of origin for ...

Biyomühendislik

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.

İletken Mürekkepler düzlemsel ve Üç Boyutlu Baskı

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

Photoacoustic Cystography

Conventional pediatric cystography, which is based on diagnostic X-ray using a radio-opaque dye, suffers from the use of harmful ionizing radiation. The risk of bladder cancers in children due to radiation exposure is more significant than many other cancers. Here we demonstrate the feasibility of nonionizing and noninvasive photoacoustic (PA) imaging of urinary bladders, referred to as photoacoustic cystography (PAC), using near-infrared (NIR) optical absorbents (i.e. methylene blue, plasmonic gold nanostructures, or single walled carbon nanotubes) as an optical-turbid tracer. We have successfully imaged a rat bladder filled with the optical absorbing agents using a dark-field confocal PAC system. After transurethral injection of the contrast agents, the rat's bladders were photoacoustically visualized by achieving significant PA signal enhancement. The accumulation was validated by spectroscopic PA imaging. Further, by using only a laser pulse energy of less than 1 mJ/cm2 (1/20 of the safety limit), our current imaging system could map the methylene-blue-filled-rat-bladder at the depth of beyond 1 cm in biological tissues in vivo. Both in vivo and ex vivo PA imaging results validate that the contrast agents were naturally excreted via urination. Thus, there is no concern regarding long-term toxic agent accumulation, which will facilitate clinical translation.

Photoacoustic cystography (PAC) has a great potential to map urinary bladders, a radiation sensitive internal organ in pediatric patients, without using any ionizing radiation or toxic contrast agent. Here we demonstrate the use of PAC for mapping urinary bladders with an injection of optical-opaque tracers in rats in vivo.

Foto akustik Sistografi

Conventional pediatric cystography, which is based on diagnostic X-ray using a radio-opaque dye, suffers from the use of harmful ionizing radiation. The ...

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.

Üç boyutlu Mürekkep Püskürtmeli Baskı Teknolojisi Kullanımı İnsan Kıkırdak Doku Fabrikasyon

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

Analysis of Cell Migration within a Three-dimensional Collagen Matrix

The ability to migrate is a hallmark of various cell types and plays a crucial role in several physiological processes, including&#160;embryonic development, wound healing, and immune responses. However, cell migration is also a key mechanism in cancer enabling these cancer cells to detach from the primary tumor to start metastatic spreading. Within the past years various cell migration assays have been developed to analyze the migratory behavior of different cell types. Because the locomotory behavior of cells markedly differs between a two-dimensional (2D) and three-dimensional (3D) environment it can be assumed that the analysis of the migration of cells that are embedded within a 3D environment would yield in more significant cell migration data. The advantage of the described 3D collagen matrix migration assay is that cells are embedded within a physiological 3D network of collagen fibers representing the major component of the extracellular matrix. Due to time-lapse video microscopy real cell migration is measured allowing the determination of several migration parameters as well as their alterations in response to pro-migratory factors or inhibitors. Various cell types could be analyzed using this technique, including lymphocytes/leukocytes, stem cells, and tumor cells. Likewise, also cell clusters or spheroids could be embedded within the collagen matrix concomitant with analysis of the emigration of single cells from the cell cluster/ spheroid into the collagen lattice. We conclude that the 3D collagen matrix migration assay is a versatile method to analyze the migration of cells within a physiological-like 3D environment.

Cell migration is a biological phenomenon that is involved in a plethora of physiological, such as wound healing and immune responses, and pathophysiological processes, like cancer. The 3D-collagen matrix migration assay is a versatile tool to analyze the migratory properties of different cell types within in a 3D physiological-like environment.

Bir Üç boyutlu Kollajen Matrix içindeki Hücre Göç Analizi

The ability to migrate is a hallmark of various cell types and plays a crucial role in several physiological processes, including&#160;embryonic ...

Engineering Three-dimensional Epithelial Tissues Embedded within Extracellular Matrix

The architecture of branched organs such as the lungs, kidneys, and mammary glands arises through the developmental process of branching morphogenesis, which is regulated by a variety of soluble and physical signals in the microenvironment. Described here is a method created to study the process of branching morphogenesis by forming engineered three-dimensional (3D) epithelial tissues of defined shape and size that are completely embedded within an extracellular matrix (ECM). This method enables the formation of arrays of identical tissues and enables the control of a variety of environmental factors, including tissue geometry, spacing, and ECM composition. This method can also be combined with widely used techniques such as traction force microscopy (TFM) to gain more information about the interactions between cells and their surrounding ECM. The protocol can be used to investigate a variety of cell and tissue processes beyond branching morphogenesis, including cancer invasion.

This manuscript describes a soft lithography-based technique to engineer uniform arrays of three-dimensional (3D) epithelial tissues of defined geometry surrounded by extracellular matrix. This method is amenable to a wide variety of cell types and experimental contexts and allows for high-throughput screening of identical replicates.

Ekstraselüler Matrix içinde Gömülü Mühendislik Üç boyutlu Epitel Dokular

The architecture of branched organs such as the lungs, kidneys, and mammary glands arises through the developmental process of branching morphogenesis, ...

Fabrication of Three-dimensional Paper-based Microfluidic Devices for Immunoassays

Paper wicks fluids autonomously due to capillary action. By patterning paper with hydrophobic barriers, the transport of fluids can be controlled and directed within a layer of paper. Moreover, stacking multiple layers of patterned paper creates sophisticated three-dimensional microfluidic networks that can support the development of analytical and bioanalytical assays. Paper-based microfluidic devices are inexpensive, portable, easy to use, and require no external equipment to operate. As a result, they hold great promise as a platform for point-of-care diagnostics. In order to properly evaluate the utility and analytical performance of paper-based devices, suitable methods must be developed to ensure their manufacture is reproducible and at a scale that is appropriate for laboratory settings. In this manuscript, a method to fabricate a general device architecture that can be used for paper-based immunoassays is described. We use a form of additive manufacturing (multi-layer lamination) to prepare devices that comprise multiple layers of patterned paper and patterned adhesive. In addition to demonstrating the proper use of these three-dimensional paper-based microfluidic devices with an immunoassay for human chorionic gonadotropin (hCG), errors in the manufacturing process that may result in device failures are discussed. We expect this approach to manufacturing paper-based devices will find broad utility in the development of analytical applications designed specifically for limited-resource settings.

We detail a method to fabricate three-dimensional paper-based microfluidic devices for use in the development of immunoassays. Our approach to device assembly is a type of multilayer, additive manufacturing. We demonstrate a sandwich immunoassay to provide representative results for these types of paper-based devices.

İmmüno için üç boyutlu Paper-based mikroakışkan Cihazlar İmalat

Paper wicks fluids autonomously due to capillary action. By patterning paper with hydrophobic barriers, the transport of fluids can be controlled and ...

Switchable Acoustic and Optical Resolution Photoacoustic Microscopy for In Vivo Small-animal Blood Vasculature Imaging

Photoacoustic microscopy (PAM) is a fast-growing invivo imaging modality that combines both optics and ultrasound, providing penetration beyond the optical mean free path (~1 mm in skin) with high resolution. By combining optical absorption contrast with the high spatial resolution of ultrasound in a single modality, this technique can penetrate deep tissues. Photoacoustic microscopy systems can have either a low acoustic resolution and probe deeply or a high optical resolution and probe shallowly. It is challenging to achieve high spatial resolution and large depth penetration with a single system. This work presents an AR-OR-PAM system capable of both high-resolution imaging at shallow depths and low-resolution deep-tissue imaging of the same sample in vivo. A lateral resolution of 4 &#181;m with 1.4 mm imaging depth using optical focusing and a lateral resolution of 45 &#181;m with 7.8 mm imaging depth using acoustic focusing were successfully demonstrated using the combined system. Here,&#160;in vivo&#160;small-animal blood vasculature imaging is performed to demonstrate its biological imaging capability.

Switchable Acoustic and Optical Resolution Photoacoustic Microscopy for In Vivo Small-animal Blood Vasculature Imaging

Here a switchable acoustic resolution (AR) and optical resolution (OR) photoacoustic microscopy (AR-OR-PAM) system capable of both high resolution imaging at shallow depth and low resolution deep tissue imaging on the same sample in vivo is demonstrated.

Için değiştirilebilir Akustik ve Optik Çözünürlük Fotoakustik Mikroskopi<emIn vivo</em&gt; Küçük Hayvan Kan Vaskülatür Görüntüleme

Photoacoustic microscopy (PAM) is a fast-growing invivo imaging modality that combines both optics and ultrasound, providing penetration beyond the ...

Characterizing Cell Migration Within Three-dimensional In Vitro Wound Environments

Currently, most in vitro models of wound healing, such as well-established scratch assays, involve studying cell migration and wound closure on two-dimensional surfaces. However, the physiological environment in which in vivo wound healing takes place is three-dimensional rather than two-dimensional. It is becoming increasingly clear that cell behavior differs greatly in two-dimensional vs. three-dimensional environments; therefore, there is a need for more physiologically relevant in vitro models for studying cell migration behaviors in wound closure. The method described herein allows for the study of cell migration in a three-dimensional model that better reflects physiological conditions than previously established two-dimensional scratch assays. The purpose of this model is to evaluate cell outgrowth via the examination of cell migration away from a spheroid body embedded within a fibrin matrix in the presence of pro- or anti-migratory factors. Using this method, cell outgrowth from the spheroid body in a three-dimensional matrix can be observed and is easily quantifiable over time via brightfield microscopy and analysis of spheroid body area. The effect of pro-migratory and/or inhibitory factors on cell migration can also be evaluated in this system. This method provides researchers with a simple method of analyzing cell migration in three-dimensional wound associated matrices in vitro, thus increasing the relevance of in vitro cell studies prior to the use of in vivo animal models.

Characterizing Cell Migration Within Three-dimensional In Vitro Wound Environments

The goal of this protocol is to evaluate the effect of pro- and anti-migratory factors on cell migration within a three-dimensional fibrin matrix.

İçinde üç boyutlu Vitro hücre göç karakterize ortamlar yara

Currently, most in vitro models of wound healing, such as well-established scratch assays, involve studying cell migration and wound closure on ...

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.

Üç boyutlu peptid iskele memeli hücrelerinde kültür

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

Methods Article

Üç boyutlu optik çözünürlük foto akustik Mikroskopi