Name: selective cell elimination from mixed 3d culture using a near infrared photoimmunotherapy technique
Uploaded: 2016-03-14T14:00:00
Description: Watch this Scientific Journal Video about Selective Cell Elimination from Mixed 3D Culture Using a Near Infrared Photoimmunotherapy Technique at JoVE.com

Это исследование было поддержано исследовательской программой Intramural Национальных институтов здравоохранения, Национальный институт рака, Центр по исследованию рака.

Примечание: Следующий протокол описывает необходимые шаги для устранения конкретных клеток с использованием БИК-шахтное. Органы управления и другие сведения о NIR-шахтное и жизнеспособность клеток может быть найден в другом месте 18. 1. Сопряжение IR700 с моноклональными антитела...

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Демонстрируется способ устранения специфического клеточного из смешанной 3D культуры клеток без ущерба для нецелевых клеток с использованием БИК-шахтное. До сих пор не существует практического метода элиминации клеток, как только будет установлено ткани или после трансплантации. Так...

Устранение конкретных клеток, не повреждая другие клетки крайне сложно, особенно в установленном ткани, и существует острая потребность в способе устранения клеток в тканевой инженерии области. В настоящее время в области регенеративной медицины, культуры тканей с использованием эмбриональных стволовых клеток (ES), плюрипотентные стволовые клетки (ЭСК) или индуцированных плюрипотентных стволовых клеток (ИПС) являются перспективными материалами 1 - 3. Хотя эта регенерации тканей является перспективным, загрязнение нежелательных клеток является серьезной проблемой. Кроме того, существует беспокойство безопасности туморогенности после трансплантации 4,5. Хотя многие исследования были сосредоточены на этих вопросах , чтобы устранить специфические клетки, особенно в регенеративной медицине 6 - 8, ни один практический метод не был разработан. Рядом с инфракрасной photoimmunotherapy (NIR-ИПН) является лечение, основанное на антитело-photoabsorber conjugatе (APC). АРС состоит из клеток-специфических моноклональных антител (монАТ) и photoabsorber, IR700. IR700 представляет собой гидрофильную производное кремнезем-фталоцианина и не вызывает фототоксичности сам по себе 9. IR700 ковалентно конъюгированный с антителом через амидные остатков в боковой цепи молекул лизина. АРС связывает молекулы-мишени на мембране клетки, а затем вызывает почти немедленное некроз клеток после воздействия NIR света при 690 нм. Во время воздействия NIR-света, клеточные мембраны разрывов , приводящих к клеточной смерти 9 - 14. NIR-PIT доказала свою эффективность с несколькими антител или фрагментов антител, в том числе анти-EGFR, анти-HER2, анти-PSMA, анти-CD25, анти-mesothelin, анти-GPC3, и анти-СЕА 15 - 21. Таким образом, БИК-PIT могут быть использованы против широкого спектра молекул-мишеней. Кроме того, NIR-PIT является хорошо контролируемое лечение, которое позволяет избирательно лечение конкретных регионов путем ограничения NIR-LIGHт облучение 18,22. Здесь мы представляем метод устранения специфического клеточного использованием БИК-шахтное из смешанных 3D культур.

<table border="0" cellpadding="0" cellspacing="0" width="1083">
	<tbody>
		<tr height="21">
			<td height="21" style="height:21px;width:424px;">IRDye 700DX Ester Infrared Dye</td>
			<td style="width:364px;">LI-COR Bioscience (Lincoln, NE, USA)</td>
			<td style="width:191px;">929-70011</td>
			<td style="width:104px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Na2HPO4</td>
			<td>SIGMA-ALDRICH (St. Louis, MO, USA)</td>
			<td>S9763</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Sephadex G25 column (PD-10)&#160;</td>
			<td>GE Healthcare (Piscataway, NJ, USA)</td>
			<td>17-0851-01</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Coomassie (bradford) Plus protein assay</td>
			<td>Thermo Fisher Scientific Inc (Waltham, MA, USA)</td>
			<td>PI-23200</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Perfecta3D 96-Well hanging Drop Plates</td>
			<td>3D Biomatrix Inc (Ann Arbor, MI, USA)</td>
			<td>HDP1096-8</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Optical power meter</td>
			<td>Thorlabs (Newton, NJ, USA)</td>
			<td>PM100</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">LED: L690-66-60</td>
			<td>Marubeni America Co. (Santa Clara, CA, USA)</td>
			<td>L690-66-60</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Vectibix (panitumumab)</td>
			<td>Amgen (Thousand Oaks, CA, USA)</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">35mm glass bottom dish, dish size 35mm, well size 10mm</td>
			<td>Cellvis (Mountain View, CA, USA)</td>
			<td>D35-10-0-N</td>
			<td></td>
		</tr>
	</tbody>
</table>

selective cell elimination from mixed 3d culture using a near infrared photoimmunotherapy technique

Recent developments in tissue engineering offer innovative solutions for many diseases. For example, tissue engineering using induced pluripotent stem cell (iPS) emerged as a new method in regenerative medicine. Although this tissue regeneration is promising, contamination with unwanted cells during tissue cultures is a major concern. Moreover, there is a safety concern regarding tumorigenicity after transplantation. Therefore, there is an urgent need for eliminating specific cells without damaging other cells that need to be protected, especially in established tissue. Here, we present a method for specific cell elimination from a mixed 3D cell culture in vitro with near infrared photoimmunotherapy (NIR-PIT) without damaging non-targeted cells. This technique enables the elimination of specific cells from mixed cell cultures or tissues.

Для того, чтобы оптически наблюдать эффект NIR-PIT, в А431 клеточной линии, которая гиперэкспрессией EGFR, была генетически модифицированной также выразить GFP и люциферазу (А431-Luc-GFP). В качестве нецелевых БИК-шахтное, клеточная линия Balb / 3T3 был оптически изменен, чтобы выразить...

Watch this Scientific Journal Video about Selective Cell Elimination from Mixed 3D Culture Using a Near Infrared Photoimmunotherapy Technique at JoVE.com

Selective Cell Elimination from Mixed 3D Culture Using a Near Infrared Photoimmunotherapy Technique

Eliminating specific cells without damaging other cells is extremely difficult, especially in established tissue, yet there is an urgent need for a cell elimination method in the tissue engineering field. Here, we present a method for specific cell elimination from a mixed 3D cell culture using near infrared photoimmunotherapy (NIR-PIT).

Селективный Cell Исключение из смешанной 3D культуры Использование ближней инфракрасной области Photoimmunotherapy Техника

Recent developments in tissue engineering offer innovative solutions for many diseases. For example, tissue engineering using induced pluripotent stem ...

Research

JoVE Journal

Bioengineering

A Simple Hanging Drop Cell Culture Protocol for Generation of 3D Spheroids

Простые висячей капли культуре клеток Протокол Генерация 3D сфероидов

Studies of cell-cell cohesion and cell-substratum adhesion have historically been performed on monolayer cultures adherent to rigid substrates. Cells ...

Rotating Cell Culture Systems for Human Cell Culture: Human Trophoblast Cells as a Model

Вращающийся культуре клеток системы по правам культуры клеток: Человеческие клетки трофобласта в качестве модели

The field of human trophoblast research aids in  understanding the complex environment established during placentation. Due to the  nature of these ...

Preparation of 3D Fibrin Scaffolds for Stem Cell Culture Applications

Подготовка 3D фибрина лесов для стволовых Приложения культуре клеток

Stem cells are found in naturally occurring 3D microenvironments in vivo, which are often referred to as the stem cell niche 1. Culturing stem cells ...

A Novel Method for Localizing Reporter Fluorescent Beads Near the Cell Culture Surface for Traction Force Microscopy

Новый метод для локализации Reporter флуоресцентные шарики Ближних культуре клеток для увеличения силы сцепления силовой микроскопии

PA gels have long been used as a platform to study cell traction forces due to ease of fabrication and the ability to tune their elastic properties. When ...

Fluorescence-quenching of a Liposomal-encapsulated Near-infrared Fluorophore as a Tool for In Vivo Optical Imaging

Fluorescence-quenching of a Liposomal-encapsulated Near-infrared Fluorophore as a Tool for In Vivo Optical Imaging

Флуоресценции тушение липосомальной инкапсулированный в ближней инфракрасной области флуорофорные как инструмент для<em&gt; В Vivo</em&gt; Оптических изображений

Optical imaging offers a wide range of diagnostic modalities and has attracted a lot of interest as a tool for biomedical imaging. Despite the enormous ...

Tunable Hydrogels from Pulmonary Extracellular Matrix for 3D Cell Culture

Настраиваемый гидрогели от легочного внеклеточной матрицы для 3D клеточных культур

Here we present a method for establishing multiple component cell culture hydrogels for in vitro lung cell culture. Beginning with healthy en bloc lung ...

Bioprinting of Cartilage and Skin Tissue Analogs Utilizing a Novel Passive Mixing Unit Technique for Bioink Precellularization

Подложке хряща и аналогов ткани кожи, используя Роман пассивного смешивания единицы техники для Bioink Precellularization

Bioprinting is a powerful technique for the rapid and reproducible fabrication of constructs for tissue engineering applications. In this study, both ...

Fabrication of Custom Agarose Wells for Cell Seeding and Tissue Ring Self-assembly Using 3D-Printed Molds

Изготовление пользовательских агарозы скважин для заполнения клеток и тканей кольцо самостоятельной сборки с использованием 3D-печатные формы

Engineered tissues are being used clinically for tissue repair and replacement, and are being developed as tools for drug screening and human disease ...

Construction of a Multilayered Mesenchymal Stem Cell Sheet with a 3D Dynamic Culture System

Строительство многослойных мезенхимальных стволовых клеток листа с системой 3D динамичной культуры

Stem cell therapy shows a promising future in regenerating injured organ and tissues, and the cell sheet technique has been developed to improve the low ...

Synthesis of Near-Infrared Emitting Gold Nanoclusters for Biological Applications

Синтез ближнего инфракрасного излучающих золотых нанокластеров для биологического применения

Over the past decade, fluorescent gold nanoclusters (AuNCs) have witnessed growing popularity in biological applications and enormous efforts have been ...