私たちは、国立科学財団（キャリアCBET-1055379）と米国立衛生研究所（1RC4EY021357、1R01EY019951）からの寛大な支援に感謝します。また、中国奨学会から魏の歌への支援を認めている。

1。システム構成<ol><li> PAOMサブシステム <ol><li>照明光源：Nd：YAGレーザ（SPOT-10-100、Elforlight株式会社、英国：20μJ/パルス、2 nsecのパルス持続時間、最大30 kHzのパルス繰り返し率）。 </li><li> 1064nmで出力レーザは、β-バリウムボレート（BBO）結晶（CasTech、サンノゼ、CA）によって532 nmに周波数を倍になります。レーザラインミラーによってさらに分割した後、532nmの光は、シングルモード光ファイバ（P1-460A-FC-5、Thorlabs）を通じて配信され、1064 nmレーザーがトリガーダイオード（DET10A、Thorlabs）によって記録されPAの信号取得。 </li><li>シングルモード光ファイバから出たレーザ光 ​​は、検流計（GM、QS-7、ナットフィールドテクノロジー）と望遠鏡の構成（F1 = 75ミリメートルとf2 = 14ミリメートル、エドモンド·オプティクス）6によって網膜上に配信されます。 </li><li>やり場の針トランスデューサ（40 MHzの中心周波数は、16 MHzの帯域幅、0.4×0.4 mm 2の能動素子のサイズ、NIHのリソースCente超音波トランスデューサ·テクノロジーズ、南カリフォルニア大学）のrは網膜から発生するPAの信号を検出するためにまぶたに接触して配置される。超音波ゲル（Sonotech）が良い音響結合のためのトランスデューサチップや動物のまぶたの間に印加される。 </li><li> PAの信号が2アンプ（ZFL-500LN +ミニ回路、5073PR、オリンパス）で増幅され、データ収集ボード（CS14200、ゲージはアプライド）によってデジタル化されています。 </li></ol></li><li> SD-OCTサブシステム <ol><li>低コヒーレンス光源：ブロードバンドスーパールミネッセントダイオード（IPSDD0804、InPhenix、中心波長：;：50nmの6 dBの帯域幅840 nm）を6μmの軸方向の分解能を決定する。 </li><li>近赤外光は50でアームとサンプルアームを参照するために分割されている×50カスタマイズしたシングルモードファイバカプラ（オズオプティクス）。 </li><li>ホットミラー（FM02、Thorlabs）によってPAOM照明光と結合した後、10月のサンプルアームは同じスキャンと配信光学wiを共有目PAOM 6。 </li><li>自作分光器は、ラインスキャンCCDカメラ（Aviiva SM2、e2v社）は、24 kHzのラインレートを可能に、SD-OCTの干渉信号を記録するために使用されます。典型的な分光器の設計は、いくつかの以前に発行された文献11から発見され、ファイバ結合SD-OCTの分光器は現在市販されていることができます。 SD-OCTの感度は90デシベルよりも良好であることが測定されます。 </li></ol></li><li> スキャニング方式 <ol><li>検流計のスキャンを高速で2次元ラスターもPAOMレーザー発射と10月分析計の信号取得の両方をトリガアナログ出力ボード（PCI-6731、ナショナルインスツルメンツ）によって制御されます。結果として、PAOMとOCTサブシステム内のデータの取得が同期されます。 </li><li> PAOMデータ取得は、フォトダイオードの記録PAOMレーザーシーケンス（1.1.2を参照）によってトリガされます。 </li><li> 3次元体積画像や2次元眼底画像（256×256 B-スキャン画像から構成され、Bごとの行）の画像をスキャンしてください。 </li></ol></li></ol> 2。システムのアライメント<ol><li> BBO結晶の周波数逓倍の効率とシングルモード光ファイバの結合効率を最大化します。 PAOM照明光を最適化する際に職員の目の保護のためのLG3ゴーグル（Thorlabs）を着用してください。 </li><li>直径2.0ミリメートルにPAOMのファイバ出力レーザをコリメート。 </li><li>同軸にPAOMとSD-OCTの複合照明光の位置を合わせます。 </li><li> 〜0.8 mWで光をプロービング〜40 NJ /パルスと、SD-OCTでPAOM励起光を設定し、どちらも6,12アイセーフであると報告されている。 </li></ol> （3）マルチモーダル生体網膜走査で <ol><li>透明ポリプロピレンボックスにラットを移し、1.5％の濃度と10分間2.0リットル/分の流量で、イソフルランの混合物（フェニックス製薬株式会社）と、通常の空気によって動物を麻酔。 </li><li> homemaで麻酔をかけたラットを拘束デ5軸調整可能自由（ 図1）と、加熱パッド（REPTIサーム、ズーム研究所、Inc。）によって〜37℃で、その体温を維持してホルダ。実験を通して1.0％の濃度と1.5リットル/分の流量で混合されたイソフルランと通常の空気の吸入ガスによる麻酔を維持している。 </li><li> 、手術用のハサミを使ってまつげをカット1％トロピカミド点眼液で生徒を拡張、0.5％塩酸テトラカイン点眼液を用いた虹彩括約筋の筋肉を麻痺させる。ラットの眼球角膜の脱水と白内障形成を防ぐために他のすべての分に人工涙液滴（Systane、アルコン·ラボラトリーズ社）を適用します。撮像中にパルスオキシメータ（8,600 V、Nonin医療、MN）が動物の心拍数、呼吸、血液酸素化を監視します。 </li><li> SD-OCTの照明光をオンにして、〜0.8mWとなるようにプローブ光を確認してください。 </li><li>ガルバノスキャンを有効にします。 SD-OCTの照射光の配信を揃えるラットの網膜上に、5軸動物ホルダーを調整することにより、関心領域（ROI）の網膜領域を識別します。 ROIが異なる研究要件に基づいて選択されるべきである間、ここに、視神経乳頭は、意図的に、視野の中心に配置されます。 </li><li>さらに最高の光学焦点に到達するまで、両方の走査方向における網膜断面（ラスタ走査方向を切り換えることによって）のSD-OCT画像品質を最適化するために、動物のホルダーを調整します。 </li><li> 5軸調整可能なプラットフォーム上で針器を準備し、トランスデューサの先端に超音波ゲルのドロップを適用し、優しく動物まぶたにトランスデューサの先端にお問い合わせください。 </li><li>外部トリガモードにPAOMレーザーを設定し、ガルバノスキャンを開始し、動物の網膜のPAOM断面画像のリアルタイム表示を有効にしてください。 PAOMイメージが最良の信号対雑音比（SNR）を有し、その間に、均等にdistを示すまで、慎重にトランスデューサの向きを調整両方の走査方向におけるributed PAの振幅パターン。 </li><li>スキャンパラメータを設定し、SD-OCT及びPAOMの同時網膜イメージングを実施しています。 SD-OCT及びPAOMオフラインの3次元画像を再構成する。当社の再建コードは、Matlabで書かれていましたし、3次元可視化は、フリーウェア（Volview、Kitware）をuing達成された。 SD-OCTの復興のためのアルゴリズム11。文献で ​​見つけることができ、PAOM再建のためのアルゴリズムは、文献に記載されています。6とRef 13。必要であれば、）-3.9）3.7の手順を繰り返します。 </li><li>実験終了後、SD-OCTプローブ光をオフに直ちにホルダーから動物を削除し、それが自然に目覚めるまで、それを暖かく保つ。回復する目のために余分な時間のために暗い環境で動物を保​​管してください。動物の麻酔と画像取得を含む全実験期間は、経験豊富なオペレーターのための30分未満です。 </li></ol>

<ol>
	<li>Kinyoun, J. L., Martin, D. C., Fujimoto, W. Y., Leonetti, D. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ophthalmoscopy+versus+fundus+photographs+for+detecting+and+grading+diabetic+retinopathy.">Ophthalmoscopy versus fundus photographs for detecting and grading diabetic retinopathy.</a> Invest. Ophthalmol. Vis. Sci. 33 (6), 1888-1893 (1992).</li><li>Schuman, J. S., Wollstein, G., Farra, T., Hertzmark, E., Aydin, A., Fujimoto, J. G., Paunescu, L. A. <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+optic+nerve+head+measurements+obtained+by+optical+coherence+tomography+and+confocal+scanning+laser+ophthalmoscopy.">Comparison of optic nerve head measurements obtained by optical coherence tomography and confocal scanning laser ophthalmoscopy.</a> Am. J. Ophthalmol. 135 (4), 504-512 (2003).</li><li>Str&#248;m, C., Sander, B., Larsen, N., Larsen, M., Lund-Andersen, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Diabetic+macular+edema+assessed+with+optical+coherence+tomography+and+stereo+fundus+photography.">Diabetic macular edema assessed with optical coherence tomography and stereo fundus photography.</a> Invest. Ophthalmol. Vis. Sci. 43 (1), 241-245 (2002).</li><li>Hu, S., 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=Three-dimensional+Optical-resolution+Photoacoustic+Microscopy.">Three-dimensional Optical-resolution Photoacoustic Microscopy.</a> J. Vis. Exp. (51), e2729 (2011).</li><li>Wang, L. 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すべての実験動物手順は、ノースウェスタン大学の施設内動物のケアと使用委員会によって承認された。

ここでは、SD-OCTと組み合わせるPAOMを用いたラットの目の in vivo網膜イメージングの同時の詳細な手順を紹介します。光散乱ベースのSD-OCTは、おそらく、網膜イメージング3のための臨床&quot;ゴールドスタンダード&quot;ですが、それが網膜に光吸収を検出するためには区別されません。新開発の6 PAOMは、網膜の光吸収特性を提供する唯一の光吸収ベース眼科画像診?...

integrated photoacoustic ophthalmoscopy and spectral-domain optical coherence tomography

臨床診断と主要な眼疾患の基本的な調査の両方が大幅に様々な非侵襲的な眼科イメージング技術の恩恵を受ける。このような眼底撮影1、共焦点走査レーザー検眼鏡（cSLO）2、光コヒーレンストモグラフィー（OCT）3などの既存の網膜画像診断法は、病気の発症や進行を監視し、新たな治療戦略の開発に重要な貢献をしています。しかし、彼らは主に網膜から後方反射光子に依存しています。結果として、通常強く網膜病態生理状態に関連付けられている網膜の光吸収特性は、従来のイメージング技術によって、アクセスできません。 光音響眼底検査（PAOM）は、高感度4-7眼の光吸収コントラストの検出を可能に新興網膜イメージングモダリティである。 PAOM NaNにosecondレーザーパルスは瞳孔を介して配信し、まぶたに付着やり場の超音波振動子で検出される光音響（PA）の信号を誘導するために後部眼を横切ってスキャンされます。なぜならヘモグロビンとメラニンの強い光吸収のため、PAOMは、非侵襲的イメージング網膜と脈絡膜の血管系を、高コントラスト6,7における網膜色素上皮（RPE）メラニンが可能です。さらに重要なことは、よく発達した分光光音響イメージング5,8に基づいて、PAOMは、糖尿病性網膜症などのいくつかのまばゆいばかりの病気9の生理学および病理学を研究する際に重要になることができ、網膜血管中のヘモグロビンの酸素飽和度をマッピングするための可能性を秘めていると加齢黄斑変性症新生血管。 また、現存する唯一の光吸収ベース眼科画像診断法であること、PAOMは十分に確立された臨床眼科イメージングTEと統合することができます6,10複数の光のコントラストに基づいて、目のより包括的な解剖学的および機能的な評価を達成するためchniques。そこで本研究では、網膜の両方の光吸収と散乱特性が明らかにされているラットの in vivo網膜イメージングで PAOMと同時に用スペクトラルドメインOCT（SD-OCT）を統合することができます。システム構成、システムのアライメントや画像取得が提示される。

図2は、SD-OCT及びPAOMの2次元眼底画像はそれぞれ、アルビノラット（A とB）と着色されたラット（CとD）で同時に取得を示しています。 SD-OCT眼底画像（ 図2Aおよび2C）で、網膜血管は、光をプロービングのヘモグロビン吸収に起因する暗い外観を持っている。網膜血管（ 図2BのRV）に加えて、PAOMが原因欠けRPEのメラニンのアルビノ...

Watch this Scientific Journal Video about Integrated Photoacoustic Ophthalmoscopy and Spectral-domain Optical Coherence Tomography at JoVE.com

Integrated Photoacoustic Ophthalmoscopy and Spectral-domain Optical Coherence Tomography

光音響眼科（PAOM）、光吸収ベースのイメージングモダリティは、現在入手可能な眼科イメージング技術に網膜の補完的な評価を提供します。我々は、ラットにおける同時マルチモーダル網膜イメージングのためのスペクトラルドメイン光コヒーレンストモグラフィー（SD-OCT）と統合PAOMの使用を報告している。

統合された光音響眼底検査とスペクトラルドメイン光コヒーレンストモグラフィー

Both the clinical diagnosis and fundamental investigation of major ocular diseases greatly benefit from various non-invasive ophthalmic imaging ...

integrated-photoacoustic-ophthalmoscopy-spectral-domain-optical

Research

JoVE Journal

Bioengineering

11.4K ビュー.  Northwestern University. 光音響眼科（PAOM）、光吸収ベースのイメージングモダリティは、現在入手可能な眼科イメージング技術に網膜の補完的な評価を提供します。我々は、ラットにおける同時マルチモーダル網膜イメージングのためのスペクトラルドメイン光コヒーレンストモグラフィー（SD-OCT）と統合PAOMの使用を報告している。

ビデオ: Integrated Photoacoustic Ophthalmoscopy and Spectral-domain Optical Coherence Tomography

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.

三次元光学分解能光音響顕微鏡

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

生物工学

Detection and Isolation of Circulating Melanoma Cells using Photoacoustic Flowmetry

Circulating tumor cells (CTCs) are those cells that have separated from a macroscopic tumor and spread through the blood and lymph systems to seed secondary tumors1,2,3. CTCs are indicators of metastatic disease and their detection in blood samples may be used to diagnose cancer and monitor a patient&prime;s response to therapy. Since CTCs are rare, comprising about one tumor cell among billions of normal blood cells in advanced cancer patients, their detection and enumeration is a difficult task. We exploit the presence of pigment in most melanoma cells to generate photoacoustic, or laser induced ultrasonic waves in a custom flow cytometer for detection of circulating melanoma cells (CMCs)4,5. This process entails separating a whole blood sample using centrifugation and obtaining the white blood cell layer. If present in whole blood, CMCs will separate with the white blood cells due to similar density. These cells are resuspended in phosphate buffered saline (PBS) and introduced into the flowmeter. Rather than a continuous flow of the blood cell suspension, we induced two phase flow in order to capture these cells for further study. In two phase flow, two immiscible liquids in a microfluidic system meet at a junction and form alternating slugs of liquid6,7. PBS suspended white blood cells and air form microliter slugs that are sequentially irradiated with laser light. The addition of a surfactant to the liquid phase allows uniform slug formation and the user can create different sized slugs by altering the flow rates of the two phases. Slugs of air and slugs of PBS with white blood cells contain no light absorbers and hence, do not produce photoacoustic waves. However, slugs of white blood cells that contain even single CMCs absorb laser light and produce high frequency acoustic waves. These slugs that generate photoacoustic waves are sequestered and collected for cytochemical staining for verification of CMCs.

We have developed a flow cytometer using laser induced ultrasound to detect circulating melanoma cells as an early indicator of metastatic disease.

光音響流量測定法を用いて循環メラノーマ細胞の検出と分離

Circulating tumor cells (CTCs) are those cells that have separated from a macroscopic tumor and spread through the blood and lymph systems to seed ...

In vitro Synthesis of Native, Fibrous Long Spacing and Segmental Long Spacing Collagen

Collagen fibrils are present in the extracellular matrix of animal tissue to provide structural scaffolding and mechanical strength. These native collagen fibrils have a characteristic banding periodicity of ~67 nm and are formed in vivo through the hierarchical assembly of Type I collagen monomers, which are 300 nm in length and 1.4 nm in diameter. In vitro, by varying the conditions to which the monomer building blocks are exposed, unique structures ranging in length scales up to 50 microns can be constructed, including not only native type fibrils, but also fibrous long spacing and segmental long spacing collagen. Herein, we present procedures for forming the three different collagen structures from a common commercially available collagen monomer. Using the protocols that we and others have published in the past to make these three types typically lead to mixtures of structures. In particular, unbanded fibrils were commonly found when making native collagen, and native fibrils were often present when making fibrous long spacing collagen. These new procedures have the advantage of producing the desired collagen fibril type almost exclusively. The formation of the desired structures is verified by imaging using an atomic force microscope.

In vitro Synthesis of Native, Fibrous Long Spacing and Segmental Long Spacing Collagen

Simple and reproducible procedures are described for making three structurally distinct collagen assemblies from a common commercially available Type I collagen monomer. Native type, fibrous long spacing or segmental long spacing collagen can be constructed by varying the conditions to which the 300 nm long and 1.4 nm diameter monomer building block is exposed.

ネイティブ、繊維状の長い間隔と分節ロング間隔コラーゲンのin vitro合成

Collagen fibrils are present in the  extracellular matrix of animal tissue to provide structural scaffolding and  mechanical strength. These native ...

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.

光音響膀胱造影

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

Preparation and Photoacoustic Analysis of Cellular Vehicles Containing Gold Nanorods

Gold nanorods are attractive for a range of biomedical applications, such as the photothermal ablation and the photoacoustic imaging of cancer, thanks to their intense optical absorbance in the near-infrared window, low cytotoxicity and potential to home into tumors. However, their delivery to tumors still remains an issue. An innovative approach consists of the exploitation of the tropism of tumor-associated macrophages that may be loaded with gold nanorods in vitro. Here, we describe the preparation and the photoacoustic inspection of cellular vehicles containing gold nanorods. PEGylated gold nanorods are modified with quaternary ammonium compounds, in order to achieve a cationic profile. On contact with murine macrophages in ordinary Petri dishes, these particles are found to undergo massive uptake into endocytic vesicles. Then these cells are embedded in biopolymeric hydrogels, which are used to verify that the stability of photoacoustic conversion of the particles is retained in their inclusion into cellular vehicles. We are confident that these results may provide new inspiration for the development of novel strategies to deliver plasmonic particles to tumors.

We show the preparation and address the feasibility of cellular vehicles containing gold nanorods for the photoacoustic imaging of cancer.

ゴールドナノロッドを含む細胞車の作製と光音響分析

Gold nanorods are attractive for a range of biomedical applications, such as the photothermal ablation and the photoacoustic imaging of cancer, thanks to ...

Computed Tomography and Optical Imaging of Osteogenesis-angiogenesis Coupling to Assess Integration of Cranial Bone Autografts and Allografts

A major parameter determining the success of a bone-grafting procedure is vascularization of the area surrounding the graft. We hypothesized that implantation of a bone autograft would induce greater bone regeneration by abundant blood vessel formation. To investigate the effect of the graft on neovascularization at the defect site, we developed a micro&#8211;computed tomography (&#181;CT) approach to characterize newly forming blood vessels, which involves systemic perfusion of the animal with a polymerizing contrast agent. This method enables detailed vascular analysis of an organ in its entirety. Additionally, blood perfusion was assessed using fluorescence imaging (FLI) of a blood-borne fluorescent agent. Bone formation was quantified by FLI using a hydroxyapatite-targeted probe and &#181;CT analysis. Stem cell recruitment was monitored by bioluminescence imaging (BLI) of transgenic mice that express luciferase under the control of the osteocalcin promoter. Here we describe and demonstrate preparation of the allograft, calvarial defect surgery, &#181;CT scanning protocols for the neovascularization study and bone formation analysis (including the in vivo perfusion of contrast agent), and the protocol for data analysis.
The 3D high-resolution analysis of vasculature demonstrated significantly greater angiogenesis in animals with implanted autografts, especially with respect to arteriole formation. Accordingly, blood perfusion was significantly higher in the autograft group by the 7th day after surgery. We observed superior bone mineralization and measured greater bone formation in animals that received autografts. Autograft implantation induced resident stem cell recruitment to the graft-host bone suture, where the cells differentiated into bone-forming cells between the 7th and 10th postoperative day. This finding means that enhanced bone formation may be attributed to the augmented vascular feeding that characterizes autograft implantation. The methods depicted may serve as an optimal tool to study bone regeneration in terms of tightly bounded bone formation and neovascularization.

Implantation of autologous and allogeneic bone grafts constitute accepted approaches to treat major craniofacial bone loss. Yet the effect of graft composition on the interplay among neovascularization, cell differentiation and bone formation is unclear. We present a multimodal imaging protocol aimed to elucidate the angiogenesis-osteogenesis interdependence at the graft proximity.

コンピュータ断層撮影法と頭蓋骨自家移植と同種移植片の統合を評価するために、骨形成、血管新生のカップリングの光イメージング

A major parameter determining the success of a bone-grafting procedure is vascularization of the area surrounding the graft. We hypothesized that ...

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.

切り替え可能な音響光学分解能光音響顕微鏡<em&gt;インビボ</em&gt;小動物の血管系イメージング

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

A High-performance Compact Photoacoustic Tomography System for In Vivo Small-animal Brain Imaging

In vivo small-animal imaging has an important role to play in preclinical studies. Photoacoustic tomography (PAT) is an emerging hybrid imaging modality that shows great potential for both preclinical and clinical applications. Conventional optical parametric oscillator-based PAT (OPO-PAT) systems are bulky and expensive and cannot provide high-speed imaging. Recently, pulsed-laser diodes (PLDs) have been successfully demonstrated as an alternative excitation source for PAT. Pulsed-laser diode PAT (PLD-PAT) has been successfully demonstrated for high-speed imaging on photoacoustic phantoms and biological tissues. This work provides a visualized experimental protocol for in vivo brain imaging using PLD-PAT. The protocol includes the compact PLD-PAT system configuration and its description, animal preparation for brain imaging, and a typical experimental procedure for 2D cross-sectional rat brain imaging. The PLD-PAT system is compact and cost-effective and can provide high-speed, high-quality imaging. Brain images collected in vivo at various scan speeds are presented.

A High-performance Compact Photoacoustic Tomography System for In Vivo Small-animal Brain Imaging

A compact pulsed-laser diode-based photoacoustic tomography (PLD-PAT) system for high-speed in vivo brain imaging in small animals is demonstrated.

高性能小型光音響トモグラフィーシステム<em&gt;インビボ</em&gt;小動物脳イメージング

In vivo small-animal imaging has an important role to play in preclinical studies. Photoacoustic tomography (PAT) is an emerging hybrid imaging modality ...

Multimodal Volumetric Retinal Imaging by Oblique Scanning Laser Ophthalmoscopy (oSLO) and Optical Coherence Tomography (OCT)

While fluorescence imaging is widely used in ophthalmology, a large field of view (FOV) three-dimensional (3D) fluorescence retinal image is still a big challenge with the state-of-the-art retinal imaging modalities because they would require z-stacking to compile a volumetric dataset. Newer optical coherence tomography (OCT) and OCT angiography (OCTA) systems overcome these restrictions to provide three-dimensional (3D) anatomical and vascular images, but the dye-free nature of OCT cannot visualize leakage indicative of vascular dysfunction. This protocol describes a novel oblique scanning laser ophthalmoscopy (oSLO) technique that provides 3D volumetric fluorescence retinal imaging. The setup of the imaging system generates the oblique scanning by a dove tail slider and aligns the final imaging system at an angle to detect fluorescent cross-sectional images. The system uses the laser scanning method, and therefore, allows an easy incorporation of OCT as a complementary volumetric structural imaging modality. In vivo imaging on rat retina is demonstrated here. Fluorescein solution is intravenously injected to produce volumetric fluorescein angiography (vFA).

Multimodal Volumetric Retinal Imaging by Oblique Scanning Laser Ophthalmoscopy oSLO and Optical Coherence Tomography OCT

Here, we present a protocol to get a large field of view (FOV) three-dimensional (3D) fluorescence and OCT retinal image by using a novel imaging multimodal platform. We will introduce the system setup, the method of alignment, and the operational protocols. In vivo imaging will be demonstrated, and representative results will be provided.

斜め走査レーザー眼底 (オスロ) と光干渉断層計 (OCT) によるマルチモーダルインタ体積網膜イメージング

While fluorescence imaging is widely used in ophthalmology, a large field of view (FOV) three-dimensional (3D) fluorescence retinal image is still a big ...

Nondestructive Monitoring of Degradable Scaffold-Based Tissue-Engineered Blood Vessel Development Using Optical Coherence Tomography

Engineered vascular grafts with structural and mechanical properties similar to natural blood vessels are expected to meet the growing demand for arterial bypass. Characterization of the growth dynamics and remodeling process of degradable polymer scaffold-based tissue-engineered blood vessels (TEBVs) with pulsatile stimulation is crucial for vascular tissue engineering. Optical imaging techniques stand out as powerful tools for monitoring vascularization of engineered tissue enabling high-resolution imaging in real-time culture. This paper demonstrates a nondestructive and fast real-time imaging strategy to monitor the growth and remodeling of TEBVs in long-term culture by using optical coherence tomography (OCT). Geometric morphology is evaluated, including vascular remodeling process, wall thickness, and comparison of TEBV thickness in different culture time points and presence of pulsatile stimulation. Finally, OCT provides practical possibilities for real-time observation of the degradation of polymer in the reconstructing tissues under pulsatile stimulation or not and in each vessel segment, by compared with the assessment of polymer degradation using scanning electron microscopic(SEM) and polarized microscope.

A step by step protocol for nondestructive and long-period monitoring the process of vascular remodeling and scaffold degradation in real-time culture of biodegradable polymeric scaffold-based tissue-engineered blood vessels with pulsatile stimulation using optical coherence tomography is described here.

光干渉断層計を用いた分解足場ベース組織設計血容器開発の非破壊モニタリング

Engineered vascular grafts with structural and mechanical properties similar to natural blood vessels are expected to meet the growing demand for arterial ...