This work is funded by the Cure for Cancer Foundation, Dutch Technology Foundation (STW) and The Netherlands Organisation for Health Research and Development (ZonMw).

sunulan prosedür Akademik Tıp Merkezi Amsterdam, kayıt numarası NL41985.018 Kurumsal Değerlendirme Kurulu tarafından onaylanmış bir araştırma protokolü altında gerçekleşir. Yazılı bilgilendirilmiş onam tüm katılımcılardan gereklidir. 1. Sistem <ol><li> Bu deney için, bir 1,280-1,350 nm dalga boyu bandı 13 çalışan, bir Fourier etki Ekim sistemini kullanın. Fourier etki düşük tutarlılık enterforemetre ilk nesil zaman etki Ekim sistemlere göre veri toplama hızını artırır, sürekli tarama sağlar. Not: Ekim sistemi 90 ° açı ~ de helezonik bir fiber optik prob ile tarama arabirim. Bu 2.7f bir dış çapa (0.9 mm) ila 135 cm arasında bir sokulabilir uzunluğa sahiptir. Sonda 54 mm geri çekilme aralığı ile bir tahrik motoru ve optik denetleyicisi (montaj yuvası) ile Ekim konsoluna bağlanır. edinilen Ekim veri kümeleri 541 kesit görüntüleri (B-taramalar) w oluşur15 um bir eksenel çözünürlük i (Şekil 1, 2). </li><li> Doğru ve tekrarlanabilir bir zayıflatma ölçümleri sağlamak için, ölçmek suretiyle kalibre μ okt Kodach ve diğ., 14, 15 ile daha önce tarif edildiği gibi (örneğin, Intralipid) bir yağ emülsiyonundan ağırlık yüzdesi, göre artan konsantrasyonlarda için. Kısacası: <ol><li> 0.125, 0.250, 0.5, 1.0, 2.0, 4.0, 10, 15 ve 20 (stok) yüzde konsantrasyonlarının elde edilmesi için mineralden arındırılmış H2 O ile% 20 yağ emülsiyonu standart toplu seyreltin. <ol><li> Yağ emülsiyonu karışımı, 200 ml Ekim duyargasını ve Ekim ölçümü kazanır. </li><li> Çapraz referans literatürde bilinen değerleri μ Ekim değerler ekstre. </li></ol></li></ol></li></ol> 2. Time Out ve Hasta Konumlandırma <ol><li> Prosedürü başlamadan önce, isim, doğum, prosedür, pr tarihini kontrol bir &quot;zaman aşımı&quot; gerçekleştirmekocedural tarafı, antikoagülan kullanımı ve alerjiler. </li><li> Tümör konumuna bağlı olarak, ya eğilimli ya da lateral dekübit pozisyonunda hastayı yerleştirin. Yeterli desteği ile hastayı sağlayın ve o / o 20 ila 40 dakikalık bir süre boyunca bu pozisyonda rahat olmasını beklediğini eğer doğrulayın. </li><li> Ultrason (US) 16 kullanılarak, tümörün lokalize ve kalıcı mürekkep ile cilt üzerinde iğne giriş noktasını işaretlemek. NOT: bilgisayarlı tomografi (BT) kullanırken, erişim iğnenin tercih konumunu lokalize esnek bir iğne rehberlik şablonu kullanın. </li></ol> 3. Dezenfeksiyon ve Steril Drapaj <ol><li> Bir cerrahi cap ve ağız kapağı üzerine koyun. </li><li> (2.3 adım) daha önce yerleştirilen iğne giriş işaretini kaldırmak için özen, bir klorheksidin / alkol solüsyonu kullanarak ponksiyon çevresindeki deriyi temizleyin. Geniş bir alanı Dezenfeksiyon beklenmedik erişim iğne yeniden konumlandırılması durumunda ek temizlik için gerekliliğini önleyecektir. </li><li> R10 ml&#39;lik bir şırınga, bir kör aspirasyon iğnesi, bir 21 G enjeksiyon iğnesi, bir neşter, bir 15 G, eş-eksenli iğnenin, bir 18 G, bir trokar iğnesini ve 16: steril içeriğin egard içeren perkütan ponksiyonu ayarlamak açmak G kor biyopsi tabancası. </li><li> Sonradan el dezenfektan uygulanması, ellerinizi iyice yıkayın. Cerrahi önlük ve steril eldiven giyin. </li><li> Steril örtüler hastayı örtün. </li><li> Ultrason probunun etrafında steril bir kapak uygulayın ve yerinde iğne kılavuzunu düzeltmek. </li></ol> 4. Ekim Hazırlık <ol><li> Ekim konsolunu başlatın ve konsol arabirimi kullanılarak hasta kimliği etiketli alanlar, soyadı, ad ve DOB (doğum tarihi) hasta ayrıntıları girin. </li><li> Steril içeriğin ilgili olarak, bir Ekim prob, steril montaj dok kapağı ve 5 ml&#39;lik luer-lock şırınga içeren Ekim paketini açmak. </li><li> Ekim konsolu montaj dock steril kapağı uygulayın. Steril olmayan montaj rıhtım gerektirir RehberlikBir asistan yardımı. </li><li> % 0.9 NaCl ile 5 ml şırınga doldurun ve yıkama noktasına takın. Su sonda kapağının distal kısmında görünene kadar Ekim probu yıkayın. </li><li> Montaj dock Ekim probu yükleyin. Döndürmek ve düzgün işleyişini doğrulayan kırmızı ışık yayacaktır prob yükledikten sonra. Kızarma ve hasar riskini en aza indirmek için yükleme sırasında koruyucu kapak probu bırakın. </li><li> Kendi kapaktan Ekim probu çıkarın. Sert bir yüzeye yerleştirin probu ve bahşiş kısaltmak için bir neşter kullanabilirsiniz. Optik fiber ve prizma baskı en aza indirmek için, kesim sırasında sondanın uzak kısmını sabitleyin. Yönlendirme için yayılan (kırmızı) ışığı kullanarak, prizmadan 5 mm distalinde kesin. </li></ol> 5. Ponksiyon <ol><li> Deri ve% 2 lidokain kullanılarak derin tabakaların (20 mg / ml) anestezisi. Etkili olması için lidokain sağlayan birkaç dakika bekleyin. Herhangi bir ağrı varsa hastayı değildir. </li><li> İğne kılavuzunu kullanma, yerGörüntüleme yoluyla konumunu doğrulamak 15 G eş-eksenli introducer iğne. Yerleştirme tatmin edici değilse, obturatörü (keskin iğne çekirdek) çıkarın. </li><li> Tümör piercing, iğnenin içinden 18 G iğne trokar yerleştirin. Yine görüntüleme ile iğnenin konumunu doğrulayın. Yerleştirme tatmin edici ise obturatörü çıkarın. </li><li> Direnç hissedene kadar trokar iğne kadar Ekim probu besleyin. </li><li> Ekim prob sabitleme ederken, tümör dokusuna Ekim prob açığa trokar iğne geri çekin. Tümör içindeki trokar iğne ucu tutulması nefes döngüleri sırasında Ekim probunun dolaşması en aza indirir. Bu prob hasar riskini düşürür. </li><li> Ekim Tarama: <ol><li> Veri kümesi başına 541 B-taramalar ayarlanmış konsol, bir Ekim tarama gerçekleştirin. Burada kullanılan Ekim sistemi belirli bir parametre ayarlaması gerektiren 5.4 sm bir uzunluk boyunca bir otomatik geri çekilme gerçekleştirir. </li><li> (Kalite, eserler için tarama ve katı doku görünümünü kontrolŞekil 1A). Pürüzler en yaygın olarak dairesel bantları, normal Ekim desen (Şekil 1B) dışarı ayakta görünür. </li><li> Eserler rescanning sonra devam ederse probu değiştirin. </li></ol></li><li> 3 Eki veri setlerinin en az kadar adımı yineleyin 5.6 elde edilir. </li><li> Yerinde sokma iğne bırakarak, Ekim prob ve trokar iğne çıkarın. </li><li> Kor biyopsi silahı kol ve görüntüleme konumunu doğrulayarak, iğnenin içinden yerleştirin. </li><li> Konumlandırma tatmin edici değilse, biyopsi tabancası ateş. </li><li> Patoloji bölümü protokolüne göre konteynere biyopsi materyali yerleştirin. Burada, kağıt dolgu ile bir petri kabı, bir yerde biyopsileri yeterli% 0.9 NaCl ile doyuruldu. </li><li> Kor biyopsi kalitesini kontrol edin ve yeterli malzeme elde edilinceye kadar adım 5.9 ve 5.10 tekrarlayın. </li></ol>

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The authors of this article have nothing to disclose.

Bu yayında biz böbrek, Ekim merkezli perkütan iğne fizibilite raporu. Bu &quot;Optik Biyopsi&quot; olarak adlandırılan tümör farklılaşması için bir klinik uygulanabilir tekniği, içine OCT gelişiminde önemli bir ilk adımdır. Bizim ilk 25 hasta kolay ve güvenli bir prosedür olarak perkütan Ekim göstermiştir. Bir optik biyopsi geleneksel çekirdek biyopsisi üzerinden iki avantajı vardır. İlk olarak, Ekim verilerin gerçek zamanlı toplama ve analiz konvansiyonel patolojinin işlem süresi 5-10 g?...

Geçtiğimiz yıl renal kitlelerin 1,2 insidansında sürekli bir artış göstermiştir. Şimdiye kadar, böbrek kitle tedavi kararları başta MR ve BT görüntüleme özellikleri, yaş ve komorbidite temelinde yapılmıştır. Ancak bu tanı yöntemleri ve klinik parametreler gerçekten bir renal kitlenin malign potansiyeli tespit incelik yoksundur. Bir kor biyopsi veya patolojik değerlendirme (teşhis) için yeterli doku ile ince iğne aspirasyon 95-100% 3 aralığında duyarlılık ve özgüllük hem objektif tümör farklılaşması sağlar. Bu nedenle biyopsi şüpheli renal kitlelerin 4,5 değerlendirilmesinde kabul görmektedir. Ancak, yeterli doku olmadan biyopsileri geciktirmek, tanı ya da böbrek parankim (non-diagnostik) Normal genel% 10-20 oranında görülür ve hatta küçük renal kitlelerde% 30&#39;a kadar olan (&lt;4 cm, KRK) kurmak Ek için sık sık gerekliliği nedeniyle tanı sürecibiyopsi 3,5. Optik koherens tomografi (OCT) renal kitle farklılaşması yukarıda belirtilen engellerin üstesinden potansiyeline sahip yeni bir görüntüleme yöntemidir. Yakın kızılötesi ışığın saçılma dayanarak, Ekim 2-3 mm etkili bir doku penetrasyonu bir 15 mikron eksenel çözünürlükte görüntüler sağlar (Şekil 1, 2). doku penetrasyonu, dokuya özel ışık saçılımı bir bileşke, her milimetresi başına sinyal yoğunluğu kaybı zayıflama katsayısı olarak ifade edilir (μ Ekim: mm-1). Faber ve diğerleri 6 tarafından tarif edildiği gibi. Histolojik özellikleri, doku farklılaşması (Şekil 3) için bir nicel parametre sağlayan Ekim değerleri μ için ilişkili olabilir. Karsinogenezis sırasında, habis hücreler artan sayıda, daha büyük ve daha düzensiz şekilli çekirdekler görüntüler<html> daha yüksek bir kırılma endeksi ve daha aktif mitokondri. Nedeniyle hücre bileşenlerinin bu aşırı ekspresyonuna bağlı olarak, μ Ekim bir değişiklik iyi huylu tümörler ya da etkilenmemiş doku 7&#39;ye habis tümörler karşılaştırırken beklenebilir. Son zamanlarda biz benign ve malign böbrek kitlelerinin 8,9 ayırt etmek için yüzeysel Ekim yeteneğini okudu. 16 hastalarda ise, tümör dokusunun intra-operatif Ekim ölçümler harici olarak yerleştirilen Ekim sonda kullanılarak elde edilmiştir. Aynı hastalarda etkilenmemiş doku Ekim ölçümleri oluşan kontrol kolu. Normal doku tümörü farklılaşması için Ekim potansiyelini teyit malign doku ile karşılaştırıldığında önemli ölçüde daha düşük medyan zayıflama katsayısı gösterdi. Bu nicel analiz gibi ürotelyal karsinom 10,11 ve vulvar epitel neoplazi farklılaşma 12 gibi malign dokusu, sınıf diğer türlerine benzer bir şekilde uygulanmıştır. ent &quot;&gt; Gerçek zamanlı görüntüleme yerinde tümör farklılaşması ile birlikte. Bu çalışmanın amacı esaslı bir perkütan, iğne tanımlamaktır sağlayan bir optik biyopsi haline Ekim geliştirmeyi hedefleyen, hastalarda Ekim yaklaşım tanısı Katı arttırıcı renal kitle. Bu yöntem açıklaması, bildiğimiz kadarıyla, ilk böbrek tümörlerinin iğne olasılığını tabanlı Ekim değerlendirmek için.

<table border="0" cellpadding="0" cellspacing="0">
	<tbody>
		<tr>
			<td>15G / 7.5cm Co-Axial Introducer Needle</td>
			<td>Angiotech, Gainesville, USA</td>
			<td>MCXS1612SX</td>
			<td></td>
		</tr>
		<tr>
			<td>18G / 20cm Trocar Needle</td>
			<td>Cook medical, Bloomington, USA</td>
			<td>DTN-18-20.0-U</td>
			<td></td>
		</tr>
		<tr>
			<td>16G / 20cm Quick-Core Biopsy Gun</td>
			<td>Cook Medical, Bloomington, USA</td>
			<td>G07827</td>
			<td></td>
		</tr>
		<tr>
			<td>Ilumien Optis PCI Optimization System (OCT &#38; FFR)</td>
			<td>St. Jude medical, St. Paul, USA</td>
			<td>C408650</td>
			<td>Part of Dragonfly Kit. 
			St. Jude medical, St. Paul, USA. (C4088643)</td>
		</tr>
		<tr>
			<td>Dragonfly Duo Imaging Catheter</td>
			<td>LightLab Imaging, Westford, USA</td>
			<td>C408644</td>
			<td>Part of Dragonfly Kit. 
			St. Jude medical, St. Paul, USA. (C4088643)</td>
		</tr>
		<tr>
			<td>Sterile Dock Cover</td>
			<td>CFI Med. Solutions, Fenton, USA</td>
			<td>200-700-00</td>
			<td>Part of Dragonfly Kit. 
			St. Jude medical, St. Paul, USA. (C4088643)</td>
		</tr>
		<tr>
			<td>5ml Luer-lock Syringe</td>
			<td>Merit Med. Syst., South Jordan, USA</td>
			<td>C408647</td>
			<td></td>
		</tr>
		<tr>
			<td>10ml Syringe</td>
			<td>BD, Franklin Lakes, USA</td>
			<td>300912</td>
			<td></td>
		</tr>
		<tr>
			<td>18G Blunt Fill Needle</td>
			<td>BD, Franklin Lakes, USA</td>
			<td>305180</td>
			<td></td>
		</tr>
		<tr>
			<td>21G Injection Needle</td>
			<td>BD, Franklin Lakes, USA</td>
			<td>301155</td>
			<td></td>
		</tr>
		<tr>
			<td>Sterile scalpel</td>
			<td>BD, Franklin Lakes, USA</td>
			<td>372611</td>
			<td></td>
		</tr>
		<tr>
			<td>NaCl 0,9% solution</td>
			<td>Braun, Melsungen AG, Germany</td>
			<td>222434</td>
			<td></td>
		</tr>
		<tr>
			<td>Lidoca&#239;ne HCl 2% (20mg/ml) solution</td>
			<td>Braun, Melsungen AG, Germany</td>
			<td>3624480</td>
			<td></td>
		</tr>
		<tr>
			<td>Sterile Ultrasound Gel, Aquasonic 100</td>
			<td>Parker Lab. Inc., Fairfield, USA</td>
			<td>GE424609</td>
			<td></td>
		</tr>
		<tr>
			<td>Sterile Ultrasound Cover</td>
			<td>Microtek Med., Alpharetta, USA</td>
			<td>PC1289EU</td>
			<td></td>
		</tr>
		<tr>
			<td>Pathology Container</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>AMIRA software package</td>
			<td colspan="2">FEI Visualization Sciences Group, Hillsboro, USA</td>
			<td>Software platform for 3D data analysis</td>
		</tr>
		<tr>
			<td>FIJI software package (open source)</td>
			<td>Open source, http://fiji.sc/Fiji</td>
			<td></td>
			<td>Open source image processing software</td>
		</tr>
	</tbody>
</table>

in vivo, percutaneous, needle based, optical coherence tomography of renal masses

Optical coherence tomography (OCT) is the optical equivalent of ultrasound imaging, based on the backscattering of near infrared light. OCT provides real time images with a 15 &#181;m axial resolution at an effective tissue penetration of 2-3 mm. Within the OCT images the loss of signal intensity per millimeter of tissue penetration, the attenuation coefficient, is calculated. The attenuation coefficient is a tissue specific property, providing a quantitative parameter for tissue differentiation. 
Until now, renal mass treatment decisions have been made primarily on the basis of MRI and CT imaging characteristics, age and comorbidity. However these parameters and diagnostic methods lack the finesse to truly detect the malignant potential of a renal mass. A successful core biopsy or fine needle aspiration provides objective tumor differentiation with both sensitivity and specificity in the range of 95-100%. However, a non-diagnostic rate of 10-20% overall, and even up to 30% in SRMs, is to be expected, delaying the diagnostic process due to the frequent necessity for additional biopsy procedures. 
We aim to develop OCT into an optical biopsy, providing real-time imaging combined with on-the-spot tumor differentiation. This publication provides a detailed step-by-step approach for percutaneous, needle based, OCT of renal masses.

İlk 25 tümörler (23 hasta) arasında, 24 Ekim başarılı prosedürleri toplam uygulandı. Bir durumda bir prob arızası bir OCT taraması elde edememe yol açtı. Tartışma bölümünde ayrıntılı olarak tarif edilmiştir (AE) meydana gelen iki yan etkiler. Hastaların genel özellikleri Tablo 1&#39;de bulunur. Ekim konsolu edinilen veri setlerinin derhal nitel analiz için gerçek zamanlı Ekim görüntüler sağlayarak yazılımı önceden yüklenmiştir. Daha fazl...

Watch this Scientific Journal Video about In Vivo, Percutaneous, Needle Based, Optical Coherence Tomography of Renal Masses at JoVE.com

In Vivo, Percutaneous, Needle Based, Optical Coherence Tomography of Renal Masses

Optical coherence tomography (OCT) is a high resolution imaging technique that allows analysis of tissue specific optical properties providing the means for tissue differentiation. We developed needle based OCT, providing real-time imaging combined with on-the-spot tumor differentiation. This publication describes a method for percutaneous, needle based OCT of renal masses.

Vivo, Perkütan, İğne Tabanlı, Böbrek Kitlelerin Optik Koherens Tomografi

Optical coherence tomography (OCT) is the optical equivalent of ultrasound imaging, based on the backscattering of near infrared light. OCT provides real ...

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JoVE Journal

Medicine

In Vivo, Percutaneous, Needle Based, Optical Coherence Tomography of Renal Masses

8.8K Görüntüleme.  Academic Medical Center. Optical coherence tomography (OCT) is a high resolution imaging technique that allows analysis of tissue specific optical properties providing the means for tissue differentiation. We developed needle based OCT, providing real-time imaging combined with on-the-spot tumor differentiation. This publication describes a method for percutaneous, needle based OCT of renal masses.

Video: In Vivo, Percutaneous, Needle Based, Optical Coherence Tomography of Renal Masses

Doppler Optical Coherence Tomography of Retinal Circulation

Noncontact retinal blood flow measurements are performed with a Fourier domain optical coherence tomography (OCT) system using a circumpapillary double circular scan (CDCS) that scans around the optic nerve head at 3.40 mm and 3.75 mm diameters. The double concentric circles are performed 6 times consecutively over 2 sec. The CDCS scan is saved with Doppler shift information from which flow can be calculated. The standard clinical protocol calls for 3 CDCS scans made with the OCT beam passing through the superonasal edge of the pupil and 3 CDCS scan through the inferonal pupil. This double-angle protocol ensures that acceptable Doppler angle is obtained on each retinal branch vessel in at least 1 scan. The CDCS scan data, a 3-dimensional volumetric OCT scan of the optic disc scan, and a color photograph of the optic disc are used together to obtain retinal blood flow measurement on an eye. We have developed a blood flow measurement software called "Doppler optical coherence tomography of retinal circulation" (DOCTORC). This semi-automated software is used to measure total retinal blood flow, vessel cross section area, and average blood velocity. The flow of each vessel is calculated from the Doppler shift in the vessel cross-sectional area and the Doppler angle between the vessel and the OCT beam. Total retinal blood flow measurement is summed from the veins around the optic disc. The results obtained at our Doppler OCT reading center showed good reproducibility between graders and methods (&lt;10%). Total retinal blood flow could be useful in the management of glaucoma, other retinal diseases, and retinal diseases. In glaucoma patients, OCT retinal blood flow measurement was highly correlated with visual field loss (R2&gt;0.57 with visual field pattern deviation). Doppler OCT is a new method to perform rapid, noncontact, and repeatable measurement of total retinal blood flow using widely available Fourier-domain OCT instrumentation. This new technology may improve the practicality of making these measurements in clinical studies and routine clinical practice.

Total retinal blood flow is measured by Doppler optical coherence tomography and semi-automated grading software.

Retina Sirkülasyon Doppler Optik Koherens Tomografi

Noncontact retinal blood flow measurements are performed with a Fourier domain optical coherence tomography (OCT) system using a circumpapillary double ...

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Near Infrared Optical Projection Tomography for Assessments of &beta;-cell Mass Distribution in Diabetes Research

By adapting OPT to include the capability of imaging in the near infrared (NIR) spectrum, we here illustrate the possibility to image larger bodies of pancreatic tissue, such as the rat pancreas, and to increase the number of channels (cell types) that may be studied in a single specimen. We further describe the implementation of a number of computational tools that provide: 1/ accurate positioning of a specimen's (in our case the pancreas) centre of mass (COM) at the axis of rotation (AR)2; 2/ improved algorithms for post-alignment tuning which prevents geometric distortions during the tomographic reconstruction2 and 3/ a protocol for intensity equalization to increase signal to noise ratios in OPT-based BCM determinations3. In addition, we describe a sample holder that minimizes the risk for unintentional movements of the specimen during image acquisition. Together, these protocols enable assessments of BCM distribution and other features, to be performed throughout the volume of intact pancreata or other organs (e.g. in studies of islet transplantation), with a resolution down to the level of individual islets of Langerhans.

Near Infrared Optical Projection Tomography for Assessments of &amp;#946;-cell Mass Distribution in Diabetes Research

We describe the adaptation of optical projection tomography (OPT)1 to imaging in the near infrared spectrum, and the implementation of a number of computational tools. These protocols enable assessments of pancreatic &beta;-cell mass (BCM) in larger specimens, increase the multichannel capacity of the technique and increase the quality of OPT data.

Diyabet Araştırma β-hücre Kütle Dağılımı değerlendirmelerinde Kızılötesi Optik Projeksiyon Tomografi civarında

By adapting OPT to include the capability of imaging in the near infrared (NIR) spectrum, we here illustrate the possibility to image larger bodies of ...

Generation and 3-Dimensional Quantitation of Arterial Lesions in Mice Using Optical Projection Tomography

The generation and analysis of vascular lesions in appropriate animal models is a cornerstone of research into cardiovascular disease, generating important information on the pathogenesis of lesion formation and the action of novel therapies. Use of atherosclerosis-prone mice, surgical methods of lesion induction, and dietary modification has dramatically improved understanding of the mechanisms that contribute to disease development and the potential of new treatments.
Classically, analysis of lesions is performed ex vivo using 2-dimensional histological techniques. This article describes application of optical projection tomography (OPT) to 3-dimensional quantitation of arterial lesions. As this technique is non-destructive, it can be used as an adjunct to standard histological and immunohistochemical analyses.
Neointimal lesions were induced by wire-insertion or ligation of the mouse femoral artery whilst atherosclerotic lesions were generated by administration of an atherogenic diet to apoE-deficient mice.
Lesions were examined using OPT imaging of autofluorescent emission followed by complementary histological and immunohistochemical analysis. OPT clearly distinguished lesions from the underlying vascular wall. Lesion size was calculated in 2-dimensional sections using planimetry, enabling calculation of lesion volume and maximal cross-sectional area. Data generated using OPT were consistent with measurements obtained using histology, confirming the accuracy of the technique and its potential as a complement (rather than alternative) to traditional methods of analysis.
This work demonstrates the potential of OPT for imaging atherosclerotic and neointimal lesions. It provides a rapid, much needed ex vivo technique for the routine 3-dimensional quantification of vascular remodelling.

Ex vivo analysis of arterial lesions from animal models of cardiovascular disease classically relies on histological and immunohistochemical techniques. These provide 2-dimensional measurements in 3-dimensional lesions. This manuscript describes the generation of arterial lesions for quantitative analysis in 3-dimensions using optical projection tomography.

Üretim ve Farelerde arteriyel Lezyonlarının 3-Boyutlu kantitasyonu Optik Projeksiyon Tomografi ile

The generation and analysis of vascular lesions in appropriate animal models is a cornerstone of research into cardiovascular disease, generating ...

In Vivo Optical Imaging of Brain Tumors and Arthritis Using Fluorescent SapC-DOPS Nanovesicles

We describe a multi-angle rotational optical imaging (MAROI) system for in vivo monitoring of physiopathological processes labeled with a fluorescent marker. Mouse models (brain tumor and arthritis) were used to evaluate the usefulness of this method. Saposin C (SapC)-dioleoylphosphatidylserine (DOPS) nanovesicles tagged with CellVue Maroon (CVM) fluorophore were administered intravenously. Animals were then placed in the rotational holder (MARS) of the in vivo imaging system. Images were acquired in 10&#176; steps over 380&#176;. A rectangular&#160;region of interest (ROI) was placed across the full image width at the model disease site. Within the ROI, and for every image, mean fluorescence intensity was computed after background subtraction. In the mouse models studied, the labeled nanovesicles were taken up in both the orthotopic and transgenic brain tumors, and in the arthritic sites (toes and ankles). Curve analysis of the multi angle image ROIs determined the angle with the highest signal. Thus, the optimal angle for imaging each disease site was characterized. The MAROI method applied to imaging of fluorescent compounds is a noninvasive, economical, and precise tool for in vivo quantitative analysis of the disease states in the described mouse models.

In Vivo Optical Imaging of Brain Tumors and Arthritis Using Fluorescent SapC-DOPS Nanovesicles

We describe a multi-angle rotational optical imaging (MAROI) system for in vivo quantitation of a fluorescent marker delivered by saposin C (SapC)-dioleoylphosphatidylserine (DOPS) nanovesicles. Employing mouse models of cancer and arthritis, we demonstrate how the MAROI signal curve analysis can be used for the precise mapping and biological characterization of disease processes.

Floresan SapC-DOPS Nanovesicles kullanma Beyin Tümörleri ve Artrit Vivo Optik Görüntüleme

We describe a multi-angle rotational optical imaging (MAROI) system for in vivo monitoring of physiopathological processes labeled with a fluorescent ...

Preparation and Respirometric Assessment of Mitochondria Isolated from Skeletal Muscle Tissue Obtained by Percutaneous Needle Biopsy

Respirometric profiling of isolated mitochondria is commonly used to investigate electron transport chain function. We describe a method for obtaining samples of human Vastus lateralis, isolating mitochondria from minimal amounts of skeletal muscle tissue, and plate based respirometric profiling using an extracellular flux (XF) analyzer. Comparison of respirometric profiles obtained using 1.0, 2.5 and 5.0 &#956;g of mitochondria indicate that 1.0 &#956;g is sufficient to measure respiration and that 5.0 &#956;g provides most consistent results based on comparison of standard errors. Western blot analysis of isolated mitochondria for mitochondrial marker COX IV and non-mitochondrial tissue marker GAPDH indicate that there is limited non-mitochondrial contamination using this protocol. The ability to study mitochondrial respirometry in as little as 20 mg of muscle tissue allows users to utilize individual biopsies for multiple study endpoints in clinical research projects.

Methods for biopsy of Vastus lateralis, preparation of purified mitochondria, and respirometric profiling are described. The use of small muscle volume makes this technique suitable for clinical research applications.

Perkütan İğne Biyopsisi ile Elde Edilen İskelet Kası Doku İzole Mitokondri hazırlanması ve Respirometrik Değerlendirmesi

Respirometric profiling of isolated mitochondria is commonly used to investigate electron transport chain function. We describe a method for obtaining ...

Transcutaneous Assessment of Renal Function in Conscious Rodents

Glomerular filtration rate (GFR) is the gold standard to assess overall kidney function. However, traditional methods to evaluate GFR are cumbersome and time-consuming. In addition, serial blood or urine samples are required, with the associated stress for the experimental animals. A recent technique significantly reduces the investment in time and resources, minimizing the invasiveness and the animal stress, but being equally valid as the traditional approaches. The method measures transcutaneously renal function. Using an optical device and the exogenous renal marker fluorescein isothiocyanate (FITC)-sinistrin, this technique is capable of measuring the elimination kinetics of the marker through the skin. With neither blood nor urine samples nor the associated laboratory assays needed, the results of the transcutaneous measurement are almost instantaneously available. The method has been already validated in different species and successfully applied in several models of renal pathology. Moreover, due to its minimally invasive characteristics, it is suitable for sequential measurements within the same animal. Here is provided a detailed protocol to carry out the transcutaneous assessment of renal function in rodents.

Determination of glomerular filtration rate (GFR) is the gold standard to assess overall kidney function. However, traditional procedures to measure this parameter are cumbersome and require a large investment of time. Here we describe a faster and minimally invasive method to determinate GFR transcutaneously.

Bilinçli Kemirgenler Böbrek Fonksiyon Transkutanöz Değerlendirmesi

Glomerular filtration rate (GFR) is the gold standard to assess overall kidney function. However, traditional methods to evaluate GFR are cumbersome and ...

Depletion of Mouse Cells from Human Tumor Xenografts Significantly Improves Downstream Analysis of Target Cells

The use of in vitro cell line models for cancer research has been a useful tool. However, it has been shown that these models fail to reliably mimic patient tumors in different assays1. Human tumor xenografts represent the gold standard with respect to tumor biology, drug discovery, and metastasis research 2-4. Tumor xenografts can be derived from different types of material like tumor cell lines, tumor tissue from primary patient tumors4 or serially transplanted tumors. When propagated in vivo, xenografted tissue is infiltrated and vascularized by cells of mouse origin. Multiple factors such as the tumor entity, the origin of xenografted material, growth rate and region of transplantation influence the composition and the amount of mouse cells present in tumor xenografts. However, even when these factors are kept constant, the degree of mouse cell contamination is highly variable.
Contaminating mouse cells significantly impair downstream analyses of human tumor xenografts. As mouse fibroblasts show high plating efficacies and proliferation rates, they tend to overgrow cultures of human tumor cells, especially slowly proliferating subpopulations. Mouse cell derived DNA, mRNA, and protein components can bias downstream gene expression analysis, next-generation sequencing, as well as proteome analysis 5. To overcome these limitations, we have developed a fast and easy method to isolate untouched human tumor cells from xenografted tumor tissue. This procedure is based on the comprehensive depletion of cells of mouse origin by combining automated tissue dissociation with the benchtop tissue dissociator and magnetic cell sorting. Here, we demonstrate that human target cells can be can be obtained with purities higher than 96% within less than 20 min independent of the tumor type.

Human tumor xenografts are vascularized and infiltrated by cells of mouse origin during the growth phase in vivo. To circumvent the bias caused by these contaminating cells during downstream analysis, we developed a method allowing for comprehensive depletion of all mouse cells by magnetic cell sorting.

İnsan Tümör ksenogratflarının gelen Fare Hücrelerinin tükenmesi Anlamlı Hedef Hücre Alt Analizi geliştirir

The use of in vitro cell line models for cancer research has been a useful tool. However, it has been shown that these models fail to reliably mimic ...

Ultrahigh Resolution Mouse Optical Coherence Tomography to Aid Intraocular Injection in Retinal Gene Therapy Research

HR-SD-OCT is utilized to monitor the progression of photoreceptor degeneration in live mouse models, assess the delivery of therapeutic agents into the subretinal space, and to evaluate toxicity and efficacy in vivo. HR-SD-OCT uses near infrared light (800-880 nm) and has optics specifically designed for the unique optics of the mouse eye with sub-2-micron axial resolution. Transgenic mouse models of outer retinal (photoreceptor) degeneration and controls were imaged to assess the disease progression. Pulled glass microneedles were used to deliver sub retinal injections of adeno-associated virus (AAV) or nanoparticles (NP) via a trans-scleral and trans-choroidal approach. Careful positioning of the needle into the subretinal space was required prior to a calibrated pressure injection, which delivers fluid into the sub retinal space. Real time subretinal surgery was conducted on our retinal imaging system (RIS). HR-SD-OCT demonstrated progressive uniform retinal degeneration due to expression of a toxic mutant human mutant rhodopsin (P347S) (RHOP347S) transgene in mice. HR-SD-OCT allows rigorous quantification of all the retinal layers. Outer nuclear layer (ONL) thickness and photoreceptor outer segment length (OSL) measurements correlate with photoreceptor vitality, degeneration, or rescue. The RIS delivery system allows real-time visualization of subretinal injections in neonatal (~P10-14) or adult mice, and HR-SD-OCT immediately determines success of delivery and maps areal extent. HR-SD-OCT is a powerful tool that can evaluate the success of subretinal surgery in mice, in addition to measuring vitality of photoreceptors in vivo. HR-SD-OCT can also be used to identify uniform animal cohorts to evaluate the extent of retinal degeneration, toxicity, and therapeutic rescue in preclinical gene therapy research studies.

Here we demonstrate a novel approach to using high resolution spectral-domain optical coherence tomography (HR-SD-OCT) to assist delivery of gene therapy agents into the subretinal space, assess its areal coverage, and characterize photoreceptor vitality.

Ultrahigh çözünürlük fare optik koherens tomografi göz içi enjeksiyon Retina gen terapisi araştırma yardımcı olmak için

HR-SD-OCT is utilized to monitor the progression of photoreceptor degeneration in live mouse models, assess the delivery of therapeutic agents into the ...

In Vivo Imaging of Cx3cr1gfp/gfp Reporter Mice with Spectral-domain Optical Coherence Tomography and Scanning Laser Ophthalmoscopy

Spectral domain optical coherence tomography (SD-OCT) and scanning laser ophthalmoscopy (SLO) are extensively used in experimental ophthalmology. In the present protocol, mice expressing green fluorescent protein (gfp) under the promoter of Cx3cr1 (BALB/c-Cx3cr1gfp/gfp) were used to image microglia cells in vivo in the retina. Microglia are resident macrophages of the retina and have been implicated in several retinal diseases1,2,3,4,5,6. This protocol provides a detailed approach for generation of retinal B-scans, with SD-OCT, and imaging of microglia cell distribution in Cx3cr1gfp/gfp mice with SLO in vivo, using an ophthalmic imaging platform system. The protocol can be used in several reporter mouse lines. However, there are some limitations to the protocol presented here. First, both SLO and SD-OCT, when used in the high-resolution mode, collect data with high axial resolution but the lateral resolution is lower (3.5 &#181;m and 6 &#181;m, respectively). Moreover, the focus and saturation level in SLO is highly dependent on parameter selection and correct alignment of the eye. Additionally, using devices designed for human patients in mice is challenging due to the higher total optical power of the mouse eye compared to the human eye; this can lead to lateral magnification inaccuracies7, which are also dependent on the magnification by the mouse lens among others. However, despite that the axial scan position is dependent upon lateral magnification, the axial SD-OCT measurements are accurate8.

In Vivo Imaging of Cx3cr1gfp/gfp Reporter Mice with Spectral-domain Optical Coherence Tomography and Scanning Laser Ophthalmoscopy

This protocol describes how high-resolution imaging techniques such as spectral domain optical coherence tomography and scanning laser ophthalmoscopy can be utilized in small rodents, using an ophthalmic imaging platform system, to obtain information on retinal thickness and microglial cell distribution, respectively.

Vivo Tayf alanlı optik Koherens tomografi ve lazer Ophthalmoscopy tarama Cx3cr1gfp/gfp muhabir farelerin görüntüleme

Spectral domain optical coherence tomography (SD-OCT) and scanning laser ophthalmoscopy (SLO) are extensively used in experimental ophthalmology. In the ...

Assessment of Kidney Function in Mouse Models of Glomerular Disease

The use of murine models to mimic human kidney disease is becoming increasingly common. Our research is focused on the assessment of glomerular function in diabetic nephropathy and podocyte-specific VEGF-A knock-out mice; therefore, this protocol describes the full kidney work-up used in our lab to assess these mouse models of glomerular disease, enabling a vast amount of information regarding kidney and glomerular function to be obtained from a single mouse. In comparison to alternative methods presented in the literature to assess glomerular function, the use of the method outlined in this paper enables the glomerular phenotype to be fully evaluated from multiple aspects. By using this method, the researcher can determine the kidney phenotype of the model and assess the mechanism as to why the phenotype develops. This vital information on the mechanism of disease is required when examining potential therapeutic avenues in these models. The methods allow for detailed functional assessment of the glomerular filtration barrier through measurement of the urinary albumin creatinine ratio and individual glomerular water permeability, as well as both structural and ultra-structural examination using the Periodic Acid Schiff stain and electron microscopy. Furthermore, analysis of the genes dysregulated at the mRNA and protein level enables mechanistic analysis of glomerular function. This protocol outlines the generic but adaptable methods that can be applied to all mouse models of glomerular disease.

This protocol describes a full kidney work-up that should be carried out in mouse models of glomerular disease. The methods allow for detailed functional, structural, and mechanistic analysis of glomerular function, which can be applied to all mouse models of glomerular disease.

Fare modelleri Glomerular hastalığı böbrek fonksiyonlarının değerlendirilmesi

The use of murine models to mimic human kidney disease is becoming increasingly common. Our research is focused on the assessment of glomerular function ...