This material is based on work supported by the National Institutes of Health (1R03-CA182052, 1R15-CA202662), the National Science Foundation Graduate Research Fellowship Program (G.G., DGE-1450079), the Arkansas Biosciences Institute, and the University of Arkansas Doctoral Academy Fellowship. Any opinions, findings, and conclusions or recommendations expressed in this material are those of the authors and do not necessarily reflect the views of the acknowledged funding agencies.

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The authors declare that they have no competing financial interests.

<html> 멀티 모달 고해상도 이미지 및 여기에보고 서브 확산 반사율 스펙트럼 섬유 다발 microendoscope 최적화 인간 또는 동물 연구 내시경 또는 휴대용 사용을 포함하여 다양한 용도 연구자에 의해 사용될 수있다. 그것은 따라서 두 개의 서로 다른 조직 깊이에서 헤모글로빈 농도, 멜라닌 농도, 조직의 산소 포화도 측정과 함께 생체 내 혀끝의 조직 마이크로 아키텍처의 시각화를위한 유?...

섬유 다발 microendoscopy 기술은 일반적으로 영상 기술 또는 분광 기술들의 조합을 사용하여 생체 조직에서 분석한다. 1-3 그러한 영상 법 고해상도 형광 microendoscopy 서브 셀룰러 해상도 이미지 수 혀끝 조직 미세 구조를 소형으로 , 마이크로 시야, 같은 proflavine, 형광, 또는 pyranine 잉크 등의 국소 조영제를 사용. 1,3-11이 영상 양상이 질적으로 저와 함께 실시간으로 질병과 건강 상피 조직을 차별화 임상 성능을 약속 보여 주었다 간 관찰자 변화. 8 때때로, 연구자는 세포 핵 크기 또는 선 곳은 양적 특징을 추출하는 고해상도 형광 현미경 데이터를 사용하지만이 조직 형태를 시각화 대상으로 주로 질적 기술 남아있다. -1,3,8- 트라이 한편 10 분광 기술, 예컨대확산 반사율 분광학으로, 기능 조직의 정보를 제공하고 정량적으로 여러 장기에 암 병변을 확인하는 임상 성능을 약속 보여 주었다 대상으로한다. 2,12-15 따라서, 잠재적으로, 또한 관찰자 간 변이성을 감소 조직 마이크로 아키텍처의 실시간 시각화를 유지하고, 조직 상태의보다 완전한 분석을 제공하는 양식의 두 가지 유형을 포함하는 장치에 대한 필요성이 존재한다. 이 목표를 달성하기 위해, 복합 프로브 기반 장비가 단일 광섬유 프로브 두 가지 양상 결합한 구성 하였다. 고해상도 형광 microendoscopy 및 서브 확산 반사율 스펙트럼을 정점 정성 고해상도의 화상 (11)이 방법의 공동 레지스터 지역 헤모글로빈 농도를 포함하는 두 개의 별개의 조직 깊이에서 양적 스펙트럼 정보 (기능성)와 조직 형태 (구조적 특성) ([HB는), 멜라닌 농도 ([멜), 및 산소 포화도 (SAO 2). 11,12,16이 특정 하위 확산 반사율 스펙트럼 양상이 제공하는 두 개의 고유 한 조직의 깊이를 샘플링하기 위해 두 개의 소스 검출기 분리 (SDS에)를 사용 기저막 및 기본 조직 기질 아래로 샘플링하여 조직의 건강에 대한 포괄적 인 그림. (11) 파이버 프로브는 약 50,000 4.5 ㎛의 직경의 광섬유 요소, 1.1 mm의 클래드 직경 1.2 mm의 전체 코팅 직경 중앙 1mm 직경 화상 섬유로 구성된다. 이미지 섬유는 220 ㎛, 클래딩 직경이 오 200 μm의 직경 섬유로 둘러싸여 있습니다. 각각 200 ㎛의 다중 모드 광섬유는 거리 화상 광섬유의 중심으로부터 864 ㎛의 중심 간 거리에 위치된다. 200 μm의 멀티 모드 섬유를 각각 25 ° 떨어져있다. &#39;소스&#39;섬유와 왼쪽 200 μm의 멀티 모드 광섬유 및 추가 번째를 사용하여은 &quot;컬렉션&quot;섬유로 200 μm의 멀티 모드 섬유를 REE,이 형상은 반드시 374 μm의 세 가지 중심 간 SDS에, 730 μm의, 1051 μm의 및 1323 μm의를 만듭니다. 섬유 팁은 섬유 상수 사이의 거리를 유지하는 원통형 금속 케이스에 동봉되어 있습니다. 통형 케이싱의 직경은 3mm이다. 광섬유 프로브 (광섬유 프로브 팁을 향해) 말단부 2 피트이다. 프로브는 4피트의 전체 길이에 대해, 추가로 2 피트이다 (계측)을 향해 기단에서 여섯 각 개개의 섬유로 분리한다. (1)는 광섬유 프로브의 표현을 나타낸다. <img alt="그림 1" src="/files/ftp_upload/54564/54564fig1.jpg" /> 그림 1 :. 광섬유 프로브 설계 광섬유 프로브는 하나의 1mm 직경의 이미지 섬유 사 (200) μm의 멀티 모드 섬유로 구성되어 있습니다. 이그림의 표시를 나타낸다 (a) 금속 단부는 SDS의 374, 730를 수득 프로브 팁에서의 섬유의 형상을 제한 캡과 최 좌측 200 μm의 다중 모드 광섬유에 대하여 (눈금 막대 ≈ 1mm)로 1,051 ㎛의, (b) 섬유는 섬유 코어 광섬유 클래딩 및 섬유 피막 (스케일 바 ≈ 1mm), (c)의 섬유가 보호 아미드 외판 (눈금 막대 ≈ 1mm)을 보여주는 금속 캡 내에 제한되고, (d ) 금속 손잡이 모든 섬유를 포함하는 하나의 블랙 케이블 (눈금 막대 ≈ 4mm) 및 (e) 상기 프로브 (눈금 막대 ≈ 4mm의 원위 팁의 화상)와 프로브의 최종 말단 팁. <a href="http://ecsource.jove.com/files/ftp_upload/54564/54564fig1large.jpg" target="_blank">이 그림의 더 큰 버전을 보려면 여기를 클릭하십시오.</a> 이 복합 장비 및 관련 TECHNI다른 결합 구조 / 기능 기술이 서로 다른 양식을 결합하여 존재하지만 가야는 단일 프로브 내에서 이러한 양식의 첫 번째 조합입니다. 예를 들어, 하이퍼 스펙트 럴 영상은 몇 가지 이름을 조직 단백질 발현 분석, (19)와 빛 간섭 단층 촬영 (OCT를) 결합 개발 된 정량적 인 헤모글로빈과 멜라닌 속성, 17, 18 및 다른 기술과 넓은 필드 영상을 결합한 제품입니다. 구강에서의 사용을위한 휴대용 프로브 하부 위장관 식도 또는 내시경 사용을 포함하여 다양한 목적에 최적화 될 수있는 일반적인 광섬유 프로브를 사용하여 컴팩트하고 쉬운 구현 계측 설정이 문서 보고서 외부 피부 배치. 11,20 이 기기의 하드웨어는 확산 반사 스펙트럼을 획득 한 후 결과 있습니다 volum를 추출하는 사용자 정의 데이터 수집 및 후 처리 코드가 모두 필요합니다[혈색소], [멜], 및 상 2를 포함하는 전자 평균 조직 생리 학적 매개 변수를 설정합니다. 사용자 정의 데이터 취득 코드 (고해상도 형광 현미경) 카메라로부터의 동시 획득 및 (확산 반사율 스펙트럼의 경우) 분광계 있도록 지어졌다. 드라이버들은 다양한 프로그래밍 언어와의 통합을 허용하는 제조 업체의 웹 사이트에서 사용할 수 있습니다. 사용자 지정 후 처리 코드는 생체 [혈색소]와 [멜] (21)의 사전 흡수 값을 가져 다음 스펙트럼의 장착 곡선을 생성하는 이전에 개발 된 비선형 최적화 피팅 프로세스를 사용합니다. 22 장착 곡선이을 최소화하여 구축 자체 원료 스펙트럼 사이 χ 2 값 피팅 곡선 및 낮은 χ 2 값. 22 코드로 조직 생체 변수 ([헤모글로빈], [멜, 상투 2)을 결정하는 단계를 포함하도록 수정 될 수있다이러한 여기에 사용되는 외인성 pyranine 잉크로 다른 발색단에서 흡수, 그래서 그 대상 생리 학적 매개 변수는 영향을받지 않습니다. 이러한 [헤모글로빈], [멜, 상투이 같은 조직 상태의 생리 지표는 치료 종양 반응보고 또는 로컬 혈관 형성 및 혈관 신생의 지표로서 사용될 수있다. 14,23을 고해상도 형광 microendoscopy 양상 포함 가이드 프로브 배치를 돕고 상피 조직의 구조 및 기능 사이의 관계의 더욱 완전한 그림으로 연구자를 제공한다. 이 기사, 건설 및 복합 microendoscope의 응용 프로그램에 설명되어 있습니다. (11)

<table border="0" cellpadding="0" cellspacing="0" width="1143">
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		</tr>
		<tr height="24">
			<td height="24" style="height:24px;width:373px;">Arduino Uno - R3</td>
			<td style="width:175px;">SparkFun Electronics</td>
			<td style="width:171px;">DEV-11021</td>
			<td>Multiple suppliers</td>
		</tr>
		<tr height="24">
			<td height="24" style="height:24px;width:373px;">Electronic Breadboard - Self-Adhesive</td>
			<td style="width:175px;">SparkFun Electronics</td>
			<td style="width:171px;">PRT-12002</td>
			<td>Multiple suppliers</td>
		</tr>
		<tr height="24">
			<td height="24" style="height:24px;width:373px;">EasyDriver - Stepper Motor Driver</td>
			<td style="width:175px;">Sparkfun Electronics</td>
			<td style="width:171px;">ROB-12779</td>
			<td></td>
		</tr>
		<tr height="24">
			<td height="24" style="height:24px;width:373px;">12V, 229 mA Power Supply</td>
			<td style="width:175px;">Phihong</td>
			<td style="width:171px;">PSM03A</td>
			<td>Multiple suppliers</td>
		</tr>
		<tr height="24">
			<td height="24" style="height:24px;width:373px;">Enhanced Sensitivity USB Spectrometer (Vis-NIR)</td>
			<td style="width:175px;">Ocean Optics</td>
			<td style="width:171px;">USB2000+VIS-NIR-ES</td>
			<td></td>
		</tr>
		<tr height="24">
			<td height="24" style="height:24px;width:373px;">550 &#181;m, 0.22 NA, SMA-SMA Fiber Patch Cable</td>
			<td style="width:175px;">Thorlabs, Inc.</td>
			<td style="width:171px;">M37L01</td>
			<td></td>
		</tr>
		<tr height="24">
			<td height="24" style="height:24px;width:373px;">Custom-Built Fiber-Optic Probe</td>
			<td style="width:175px;">Myriad Fiber Imaging</td>
			<td style="width:171px;">N/A</td>
			<td></td>
		</tr>
		<tr height="24">
			<td height="24" style="height:24px;width:373px;">20% Spectralon Diffuse Reflectance Standard</td>
			<td style="width:175px;">Labsphere, Inc.</td>
			<td style="width:171px;">SRS-20-010</td>
			<td></td>
		</tr>
		<tr height="24">
			<td height="24" style="height:24px;width:373px;">Standard Yellow Highlighter</td>
			<td style="width:175px;">Sharpie</td>
			<td style="width:171px;">25005</td>
			<td>Multiple suppliers, proflavine or fluorescein can be substituted</td>
		</tr>
	</tbody>
</table>

multimodal imaging and spectroscopy fiber-bundle microendoscopy platform for non-invasive, in vivo tissue analysis

최근 섬유 다발 microendoscopy 기술 이미징 기술 또는 분광 기술들의 조합을 사용하여 생체 내 조직의 비파괴 분석을 가능하게한다. 하나의 광학 탐침으로 이미징 분광기 기술을 결합하여 조직 상태의보다 완전한 분석을 제공 할 수있다. 이 기사에서는 두 개의 서로 다른 양식은 하나의 광학 프로브로, 고해상도 형광 microendoscopy 이미징 및 확산 반사율 분광학을 결합됩니다. 고해상도 형광 microendoscopy 촬상 주로 질적 기술되지만, 종양 및 비 종양 조직 간의 효율적인 실시간 분화를 보여 주었다, 정점 조직 미세 구조를 시각화하기 위해 사용되는 기술이며. 확산 반사율 스펙트럼은 로컬 헤모글로빈 농도 멜라닌 농도, 산소 포화도를 포함한 조직 생리 학적 파라미터를 추출 할 수있는 기술이다. 이 글은 사양 r에 대해 설명계측을 구축하고 생체 인간의 피부에 기술을 보여줍니다하는 방법, 광섬유 프로브를 구성하는 equired. 이 작품은 조직 마이크로 아키텍처, 특히 혀끝의 피부 각질 세포가, 그와 관련된 생리 학적 매개 변수와 함께 공동 등록 할 수 있습니다 것으로 나타났습니다. 여기에 제시된 수단 및 섬유 다발 프로브 장기 다양한 시스템에서 사용하기위한 핸드 헬드 또는 내시경 호환 장치 중 하나로 최적화 될 수있다. 추가의 임상 연구는 상이한 상피 질환 상태에 대한 이러한 기술의 가능성을 테스트하기 위해 필요하다.

이 프로토콜에 따라, 연구자는 뷰의 전체 필드 (도 5)로 조직 부위의 포커스에 고해상도 이미지를 얻을 것이다. 표준 노란색 형광펜에서 pyranine 잉크로 염색하는 경우 등 proflavine 등의 염료로 염색하는 경우 각각의 세포 핵을 볼 수있는 반면, 세포의 개요는, 볼 수 있습니다. 스펙트럼 수집 후, 후 처리 소프트웨어 서브 확산 반사율 스펙트럼을 맞고 [헤모글로빈...

Watch this Scientific Journal Video about Multimodal Imaging and Spectroscopy Fiber-bundle Microendoscopy Platform for Non-invasive, In Vivo Tissue Analysis at JoVE.com

Multimodal Imaging and Spectroscopy Fiber-bundle Microendoscopy Platform for Non-invasive, In Vivo Tissue Analysis

The assembly and use of a multimodal microendoscope is described which can co-register superficial tissue image data with tissue physiological parameters including hemoglobin concentration, melanin concentration, and oxygen saturation. This technique can be useful for evaluating tissue structure and perfusion, and can be optimized for individual needs of the investigator.

멀티 모달 이미징 및 비 침습적에 대한 분광학 섬유 다발 Microendoscopy 플랫폼,<em&gt; 생체</em&gt; 조직 분석

Recent fiber-bundle microendoscopy techniques enable non-invasive analysis of in vivo tissue using either imaging techniques or a combination of ...

multimodal-imaging-spectroscopy-fiber-bundle-microendoscopy-platform

Research

JoVE Journal

Bioengineering

Multimodal Imaging and Spectroscopy Fiber-bundle Microendoscopy Platform for Non-invasive, In Vivo Tissue Analysis

7.8K 조회수.  University of Arkansas. The assembly and use of a multimodal microendoscope is described which can co-register superficial tissue image data with tissue physiological parameters including hemoglobin concentration, melanin concentration, and oxygen saturation. This technique can be useful for evaluating tissue structure and perfusion, and can be optimized for individual needs of the investigator.

비디오: Multimodal Imaging and Spectroscopy Fiber-bundle Microendoscopy Platform for Non-invasive, In Vivo Tissue Analysis

High-resolution Fiber-optic Microendoscopy for in situ Cellular Imaging

Many biological and clinical studies require the longitudinal study and analysis of morphology and function with cellular level resolution. Traditionally, multiple experiments are run in parallel, with individual samples removed from the study at sequential time points for evaluation by light microscopy. Several intravital techniques have been developed, with confocal, multiphoton, and second harmonic microscopy all demonstrating their ability to be used for imaging in situ 1. With these systems, however, the required infrastructure is complex and expensive, involving scanning laser systems and complex light sources. Here we present a protocol for the design and assembly of a high-resolution microendoscope which can be built in a day using off-the-shelf components for under US$5,000. The platform offers flexibility in terms of image resolution, field-of-view, and operating wavelength, and we describe how these parameters can be easily modified to meet the specific needs of the end user.
We and others have explored the use of the high-resolution microendoscope (HRME) in in vitro cell culture 2-5, in excised 6 and living animal tissues 2,5, and in human tissues in vivo 2,7. Users have reported the use of several different fluorescent contrast agents, including proflavine 2-4, benzoporphyrin-derivative monoacid ring A (BPD-MA) 5, and fluoroscein 6,7, all of which have received full, or investigational approval from the FDA for use in human subjects. High-resolution microendoscopy, in the form described here, may appeal to a wide range of researchers working in the basic and clinical sciences. The technique offers an effective and economical approach which complements traditional benchtop microscopy, by enabling the user to perform high-resolution, longitudinal imaging in situ.

High-resolution Fiber-optic Microendoscopy for in situ Cellular Imaging

In many biological and clinical situations it is advantageous to study cellular processes as they evolve in their native microenvironment. Here we describe the assembly and use of a low-cost fiber-optic microscope which can provide real time imaging in cell culture, animal studies, and clinical patient studies.

고해상도 광섬유 Microendoscopy에 대한 현장에서 세포 이미징

Many biological and clinical studies require the longitudinal study and analysis of morphology and function with cellular level resolution.  ...

연구

생체공학

Video-rate Scanning Confocal Microscopy and Microendoscopy

Confocal microscopy has become an invaluable tool in biology and the biomedical sciences, enabling rapid, high-sensitivity, and high-resolution optical sectioning of complex systems. Confocal microscopy is routinely used, for example, to study specific cellular targets1, monitor dynamics in living cells2-4, and visualize the three dimensional evolution of entire organisms5,6. Extensions of confocal imaging systems, such as confocal microendoscopes, allow for high-resolution imaging in vivo7 and are currently being applied to disease imaging and diagnosis in clinical settings8,9.
Confocal microscopy provides three-dimensional resolution by creating so-called "optical sections" using straightforward geometrical optics. In a standard wide-field microscope, fluorescence generated from a sample is collected by an objective lens and relayed directly to a detector. While acceptable for imaging thin samples, thick samples become blurred by fluorescence generated above and below the objective focal plane. In contrast, confocal microscopy enables virtual, optical sectioning of samples, rejecting out-of-focus light to build high resolution three-dimensional representations of samples.
Confocal microscopes achieve this feat by using a confocal aperture in the detection beam path. The fluorescence collected from a sample by the objective is relayed back through the scanning mirrors and through the primary dichroic mirror, a mirror carefully selected to reflect shorter wavelengths such as the laser excitation beam while passing the longer, Stokes-shifted fluorescence emission. This long-wavelength fluorescence signal is then passed to a pair of lenses on either side of a pinhole that is positioned at a plane exactly conjugate with the focal plane of the objective lens. Photons collected from the focal volume of the object are collimated by the objective lens and are focused by the confocal lenses through the pinhole. Fluorescence generated above or below the focal plane will therefore not be collimated properly, and will not pass through the confocal pinhole1, creating an optical section in which only light from the microscope focus is visible. (Fig 1). Thus the pinhole effectively acts as a virtual aperture in the focal plane, confining the detected emission to only one limited spatial location.
Modern commercial confocal microscopes offer users fully automated operation, making formerly complex imaging procedures relatively straightforward and accessible. Despite the flexibility and power of these systems, commercial confocal microscopes are not well suited for all confocal imaging tasks, such as many in vivo imaging applications. Without the ability to create customized imaging systems to meet their needs, important experiments can remain out of reach to many scientists.
In this article, we provide a step-by-step method for the complete construction of a custom, video-rate confocal imaging system from basic components. The upright microscope will be constructed using a resonant galvanometric mirror to provide the fast scanning axis, while a standard speed resonant galvanometric mirror will scan the slow axis. To create a precise scanned beam in the objective lens focus, these mirrors will be positioned at the so-called telecentric planes using four relay lenses. Confocal detection will be accomplished using a standard, off-the-shelf photomultiplier tube (PMT), and the images will be captured and displayed using a Matrox framegrabber card and the included software.

The complete construction of a custom, real-time confocal scanning imaging system is described. This system, which can be readily used for video-rate microscopy and microendoscopy, allows for an array of imaging geometries and applications not accessible using standard commercial confocal systems, at a fraction of the cost.

비디오 - 속도 검색 공촛점 현미경과 Microendoscopy

Confocal microscopy has become an invaluable tool in biology and the biomedical sciences, enabling rapid, high-sensitivity, and high-resolution optical ...

Non-contact, Label-free Monitoring of Cells and Extracellular Matrix using Raman Spectroscopy

Non-destructive, non-contact and label-free technologies to monitor cell and tissue cultures are needed in the field of biomedical research.1-5 However, currently available routine methods require processing steps and alter sample integrity. Raman spectroscopy is a fast method that enables the measurement of biological samples without the need for further processing steps. This laser-based technology detects the inelastic scattering of monochromatic light.6 As every chemical vibration is assigned to a specific Raman band (wavenumber in cm-1), each biological sample features a typical spectral pattern due to their inherent biochemical composition.7-9 Within Raman spectra, the peak intensities correlate with the amount of the present molecular bonds.1 Similarities and differences of the spectral data sets can be detected by employing a multivariate analysis (e.g. principal component analysis (PCA)).10
Here, we perform Raman spectroscopy of living cells and native tissues. Cells are either seeded on glass bottom dishes or kept in suspension under normal cell culture conditions (37 &deg;C, 5% CO2) before measurement. Native tissues are dissected and stored in phosphate buffered saline (PBS) at 4 &deg;C prior measurements. Depending on our experimental set up, we then either focused on the cell nucleus or extracellular matrix (ECM) proteins such as elastin and collagen. For all studies, a minimum of 30 cells or 30 random points of interest within the ECM are measured. Data processing steps included background subtraction and normalization.

Raman spectroscopy is a suitable technique for the non-contact, label-free analysis of living cells, tissue-engineered constructs and native tissues. Source-specific spectral fingerprints can be generated and analyzed using multivariate analysis.

라만 분광학을 사용하여 세포와 세포외 기질의 비 접촉, 레이블이없는 모니터링

Non-destructive, non-contact and label-free technologies to monitor cell and tissue cultures are needed in the field of biomedical research.1-5 However, ...

Viral Nanoparticles for In vivo Tumor Imaging

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

Viral Nanoparticles for In vivo Tumor Imaging

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

에 대한 바이러스 나노 입자<em&gt; 생체에</em&gt; 종양 이미징

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

Self-reporting Scaffolds for 3-Dimensional Cell Culture

Culturing cells in 3D on appropriate scaffolds is thought to better mimic the in vivo microenvironment and increase cell-cell interactions. The resulting 3D cellular construct can often be more relevant to studying the molecular events and cell-cell interactions than similar experiments studied in 2D. To create effective 3D cultures with high cell viability throughout the scaffold the culture conditions such as oxygen and pH need to be carefully controlled as gradients in analyte concentration can exist throughout the 3D construct. Here we describe the methods of preparing biocompatible pH responsive sol-gel nanosensors and their incorporation into poly(lactic-co-glycolic acid) (PLGA) electrospun scaffolds along with their subsequent preparation for the culture of mammalian cells. The pH responsive scaffolds can be used as tools to determine microenvironmental pH within a 3D cellular construct. Furthermore, we detail the delivery of pH responsive nanosensors to the intracellular environment of mammalian cells whose growth was supported by electrospun PLGA scaffolds. The cytoplasmic location of the pH responsive nanosensors can be utilized to monitor intracellular pH (pHi) during ongoing experimentation.

Biocompatible pH responsive sol-gel nanosensors can be incorporated into poly(lactic-co-glycolic acid) (PLGA) electrospun scaffolds. The produced self-reporting scaffolds can be used for in&#160;situ monitoring of microenvironmental conditions whilst culturing cells upon the scaffold. This is beneficial as the 3D cellular construct can be monitored in real-time without disturbing the experiment.

3 차원 세포 배양을위한 자기보고 발판

Culturing cells in 3D on appropriate scaffolds is thought to better mimic the in vivo microenvironment and increase cell-cell interactions. The resulting ...

A Novel Stretching Platform for Applications in Cell and Tissue Mechanobiology

Tools that allow the application of mechanical forces to cells and tissues or that can quantify the mechanical properties of biological tissues have contributed dramatically to the understanding of basic mechanobiology. These techniques have been extensively used to demonstrate how the onset and progression of various diseases are heavily influenced by mechanical cues. This article presents a multi-functional biaxial stretching (BAXS) platform that can either mechanically stimulate single cells or quantify the mechanical stiffness of tissues. The BAXS platform consists of four voice coil motors that can be controlled independently. Single cells can be cultured on a flexible substrate that can be attached to the motors allowing one to expose the cells to complex, dynamic, and spatially varying strain fields. Conversely, by incorporating a force load cell, one can also quantify the mechanical properties of primary tissues as they are exposed to deformation cycles. In both cases, a proper set of clamps must be designed and mounted to the BAXS platform motors in order to firmly hold the flexible substrate or the tissue of interest. The BAXS platform can be mounted on an inverted microscope to perform simultaneous transmitted light and/or fluorescence imaging to examine the structural or biochemical response of the sample during stretching experiments. This article provides experimental details of the design and usage of the BAXS platform and presents results for single cell and whole tissue studies. The BAXS platform was used to measure the deformation of nuclei in single mouse myoblast cells in response to substrate strain and to measure the stiffness of isolated mouse aortas. The BAXS platform is a versatile tool that can be combined with various optical microscopies in order to provide novel mechanobiological insights at the sub-cellular, cellular and whole tissue levels.

We present in this article a novel stretching platform that can be used to investigate single cell responses to complex anisotropic biaxial mechanical deformation and quantify the mechanical properties of biological tissues.

세포와 조직 Mechanobiology의 응용 프로그램에 대한 새로운 스트레칭 플랫폼

Tools that allow the application of mechanical forces to cells and tissues or that can quantify the mechanical properties of biological tissues have ...

Biofunctionalized Prussian Blue Nanoparticles for Multimodal Molecular Imaging Applications

Multimodal, molecular imaging allows the visualization of biological processes at cellular, subcellular, and molecular-level resolutions using multiple, complementary imaging techniques. These imaging agents facilitate the real-time assessment of pathways and mechanisms in vivo, which enhance both diagnostic and therapeutic efficacy. This article presents the protocol for the synthesis of biofunctionalized Prussian blue nanoparticles (PB NPs) - a novel class of agents for use in multimodal, molecular imaging applications. The imaging modalities incorporated in the nanoparticles, fluorescence imaging and magnetic resonance imaging (MRI), have complementary features. The PB NPs possess a core-shell design where gadolinium and manganese ions incorporated within the interstitial spaces of the PB lattice generate MRI contrast, both in T1 and T2-weighted sequences. The PB NPs are coated with fluorescent avidin using electrostatic self-assembly, which enables fluorescence imaging. The avidin-coated nanoparticles are modified with biotinylated ligands that confer molecular targeting capabilities to the nanoparticles. The stability and toxicity of the nanoparticles are measured, as well as their MRI relaxivities. The multimodal, molecular imaging capabilities of these biofunctionalized PB NPs are then demonstrated by using them for fluorescence imaging and molecular MRI in vitro.

This protocol describes the synthesis of biofunctionalized Prussian blue nanoparticles and their use as multimodal, molecular imaging agents. The nanoparticles have a core-shell design where gadolinium or manganese ions within the nanoparticle core generate MRI contrast. The biofunctional shell contains fluorophores for fluorescence imaging and targeting ligands for molecular targeting.

멀티 모달 분자 이미징 애플리케이션을위한 Biofunctionalized 프 러시안 블루 나노 입자

Multimodal, molecular imaging allows the visualization of biological processes at cellular, subcellular, and molecular-level resolutions using multiple, ...

Atomic Force Microscopy Imaging and Force Spectroscopy of Supported Lipid Bilayers

Atomic force microscopy (AFM) is a versatile, high-resolution imaging technique that allows visualization of biological membranes. It has sufficient magnification to examine membrane substructures and even individual molecules. AFM can act as a force probe to measure interactions and mechanical properties of membranes. Supported lipid bilayers are conventionally used as membrane models in AFM studies. In this protocol, we demonstrate how to prepare supported bilayers and characterize their structure and mechanical properties using AFM. These include bilayer thickness and breakthrough force. 
The information provided by AFM imaging and force spectroscopy help define mechanical and chemical properties of membranes. These properties play an important role in cellular processes such as maintaining cell hemostasis from environmental stress, bringing membrane proteins together, and stabilizing protein complexes.

We describe a protocol for preparation of supported lipid bilayers and its characterization using atomic force microscopy and force spectroscopy.

원자 힘 현미경 이미징 및 지원 지질 이중층의 힘 분광학

Atomic force microscopy (AFM) is a versatile, high-resolution imaging technique that allows visualization of biological membranes. It has sufficient ...

A Microfluidic Platform for High-throughput Single-cell Isolation and Culture

Studying the heterogeneity of single cells is crucial for many biological questions, but is technically difficult. Thus, there is a need for a simple, yet high-throughput, method to perform single-cell culture experiments. Here, we report a microfluidic chip-based strategy for high-efficiency single-cell isolation (~77%) and demonstrate its capability of performing long-term single-cell culture (up to 7 d) and cellular heterogeneity analysis using clonogenic assay. These applications were demonstrated with KT98 mouse neural stem cells, and A549 and MDA-MB-435 human cancer cells. High single-cell isolation efficiency and long-term culture capability are achieved by using different sizes of microwells on the top and bottom of the microfluidic channel. The small microwell array is designed for precisely isolating single-cells, and the large microwell array is used for single-cell clonal culture in the microfluidic chip. This microfluidic platform constitutes an attractive approach for single-cell culture applications, due to its flexibility of adjustable cell culture spaces for different culture strategies, without decreasing isolation efficiency.

Here, we present a protocol for isolating and culturing single cells with a microfluidic platform, which utilizes a new microwell design concept to allow for high-efficiency single cell isolation and long-term clonal culture.

높은 처리량 단일 세포의 분리 및 문화를위한 미세 유체 플랫폼

Studying the heterogeneity of single cells is crucial for many biological questions, but is technically difficult. Thus, there is a need for a simple, yet ...

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

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

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

휴대용 임상 Photoacoustic 실시간 비-침략 적 작은 동물 이미징을 위한 이미징 시스템

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

멀티 모달 이미징 및 비 침습적에 대한 분광학 섬유 다발 Microendoscopy 플랫폼,

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