The financial support of the Max-Planck Society, the Turkish Ministry of National Education (PhD fellowship to S. G.), and the German Exchange Academic Service (DAAD, PhD fellowship to T. S.) are gratefully acknowledged.

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The authors have nothing to disclose.

덴드리머 MRI 조영제의 제조 모노머 유닛의 적절한 선택을 요구한다 (즉, 하나님에 대한 킬 레이터 (III)). 그들은이 상자성 이온의 독성을 감소시키고, 현재까지 비 환식의 다양한 및 거대 고리 킬 레이터는 이러한 목적으로 제공 1-3. 이들 중, 거대 고리 형 DOTA 킬레이트는 높은 열역학적 안정성 및 불활성 운동을 보유하고, 따라서, 불활성 MRI 조영제 1,18의 제조에 가장 바람직?...

자기 공명 영상 (MRI)은 인해 비침 특성이 우수한 고유 연조직 콘트라스트 널리 생물 의학 연구 및 임상 진단에 사용하는 강력한 비 이온화 이미징 기술이다. 가장 일반적으로 사용되는 MRI 방법은 수분 신호의 밀도의 차이에 기초하여, 조직 내에서 고해상도 이미지 및 상세 정보의 제공 물 양자로부터 얻어지는 신호를 이용한다. 신호 강도 및 MRI 실험의 특이성은 또한 조영제 (CAS)를 사용하여 개선 될 수있다. 이러한 상자성 또는 초상 자성 길이 (T 1)와 횡 방향에 영향을 종 (T 2) 완화 시간은 각각 1,2이다. 폴리 카르 복실 산 리간드 란탄 족 이온의 가돌리늄 착물은 가장 일반적으로 사용되는 하나의 CA T이다. 가돌리늄 (III)는 T 1 휴식을 단축물 양성자의 시간, 따라서 MRI 실험 3에서의 신호의 콘트라스트를 증가시킨다. 그러나, 이온 성 가돌리늄은 독성; 크기는 (II) 칼슘의 근사하고 심각 칼슘 보조 세포에서 신호에 영향을줍니다. 따라서, 비 환상 및 마크로 킬레이트이 독성을 중화시키기 위해 사용된다. 다양한 여러 자리 리간드는 높은 열역학적 안정성 운동 불활성 1 가돌리늄 (III) 착체의 결과, 지금까지 개발되어왔다. 특히 그 테트라 파생 DOTA (1,4,7,10-tetraazacyclododecane-1,4,7,10-테트라)에서 12 원 azamacrocycle cyclen에 따라 사람들은,이 CA 클래스의 대부분의 연구와 적용 단지입니다. 그럼에도 GdDOTA 형 CA는 낮은 콘트라스트 효율과 빠른 신장 배출 특정 단점을 표시 저분자 시스템이다. 고분자와 다가 CA는 이러한 문제 4에 좋은 해결책이 될 수 있습니다. CA의 biodistribu 이후기 주로 자신의 크기에 의해 결정되고, 고분자 CA는 조직 내에서 더 이상 유지 시간을 표시합니다. 마찬가지로 중요한 것은, 실질적으로 획득 된 MR 신호 및 측정 품질을 향상시키는 단량체 MR 프로브 (예 GdDOTA 착체)의 증가 된 국소 농도 이러한 에이전트 결과 multivalency. 덴드리머는 MRI 4,5에 대한 다가 CA의 준비를위한 가장 바람직한 발판 사이에 있습니다. 잘 정의 된 크기의이 매우 분기 거대 분자는 표면에 다양한 커플 링 반응하는 경향이 있습니다. 이 작품에서 우리는 한 세대 4 (G4) 폴리 (아미도 아민) GdDOTA 같은 킬레이트 (DCA)에 결합 (PAMAM) 덴드리머로 구성된 MRI에 대한 덴드리머 CA의 제조, 정제 및 특성을보고합니다. 우리는 반응성 DOTA 유도체의 합성 및 PAMAM 덴드리머과의 커플 링을 설명합니다. 하나님 (III), 표준 물리 화학적 특성과 착화시 proceduDCA 재 수행 하였다. 마지막으로, MRI 실험 저분자 CA에 의한 것보다 더 강한 콘트라스트 MR 화상을 생성하기 DCA의 능력을 입증 하였다.

<table border="0" cellpadding="0" cellspacing="0" width="921">
	<tbody>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">Cyclen</td>
			<td style="width:275px;">CheMatech</td>
			<td style="width:136px;">C002</td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">tert-Butyl bromoacetate&#160;</td>
			<td style="width:275px;">Alfa Aesar</td>
			<td style="width:136px;">A14917</td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">N,N-Dimethylformamide</td>
			<td style="width:275px;">Fluka</td>
			<td style="width:136px;">40248</td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">Potassium carbonate</td>
			<td style="width:275px;">Sigma-Aldrich</td>
			<td style="width:136px;">209619</td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">4-(4-Nitrophenyl)butryic acid</td>
			<td style="width:275px;">Aldrich</td>
			<td style="width:136px;">335339</td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:251px;">Thionyl chloride&#160;</td>
			<td style="width:275px;">Acros Organics</td>
			<td style="width:136px;">382662500</td>
			<td style="width:259px;">Note: Corrosive substance; toxic if inhaled</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:251px;">Bromine</td>
			<td style="width:275px;">Acros Organics</td>
			<td style="width:136px;">402841000</td>
			<td style="width:259px;">Note: causes severe skin burns, fatal if inhaled&#160;</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">Diethyl ether</td>
			<td style="width:275px;">any source</td>
			<td style="width:136px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">Sodium sulphate</td>
			<td style="width:275px;">Acros Organics</td>
			<td style="width:136px;">196640010</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">Chloroform&#160;</td>
			<td style="width:275px;">VWR Chemicals</td>
			<td style="width:136px;">22711.29</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:251px;">tert-Butyl 2,2,2-trichloroacetimidate</td>
			<td style="width:275px;">Aldrich</td>
			<td style="width:136px;">364789</td>
			<td style="width:259px;">Note: flammable substance; irritrant to skin and eyes</td>
		</tr>
		<tr height="67">
			<td height="67" style="height:67px;width:251px;">Boron trifluoride etherate</td>
			<td style="width:275px;">Acros Organics</td>
			<td style="width:136px;">174560250</td>
			<td style="width:259px;">48 % BF3. Note: Flammable substance; causes skin burns, fatal if inhaled&#160;</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">Sodium bicarbonate</td>
			<td style="width:275px;">Acros Organics</td>
			<td style="width:136px;">424270010</td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">Ethyl-acetate</td>
			<td style="width:275px;">any source</td>
			<td style="width:136px;"></td>
			<td>For column chromatography</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">n-Hexane</td>
			<td style="width:275px;">any source</td>
			<td style="width:136px;"></td>
			<td>For column chromatography</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:251px;">Bulb-to-bulb (Kugelrohr) distillation apparatus</td>
			<td style="width:275px;">B&#252;chi</td>
			<td style="width:136px;"></td>
			<td>Model type: Glass oven B-585</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">Silicagel</td>
			<td style="width:275px;">Carl Roth GmbH</td>
			<td style="width:136px;">P090.2</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">Methanol</td>
			<td style="width:275px;">any source</td>
			<td style="width:136px;"></td>
			<td>For column chromatography</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">Dichloromethane&#160;</td>
			<td style="width:275px;">any source</td>
			<td style="width:136px;"></td>
			<td>For column chromatography</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">Ethanol</td>
			<td style="width:275px;">VWR Chemicals</td>
			<td style="width:136px;">20821.296</td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">Ammonia</td>
			<td style="width:275px;">Acros Organics</td>
			<td style="width:136px;">428381000</td>
			<td style="width:259px;">7N Solution in Methanol</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">Palladium&#160;</td>
			<td style="width:275px;">Aldrich</td>
			<td style="width:136px;">643181</td>
			<td style="width:259px;">15 % wet</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">Hydrogenation apparatus PARR</td>
			<td style="width:275px;">PARR Instrument Company</td>
			<td style="width:136px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">Celite 503</td>
			<td style="width:275px;">Aldrich</td>
			<td style="width:136px;">22151</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">Sintered glass funnel</td>
			<td style="width:275px;">any source</td>
			<td style="width:136px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">Thiophosgen</td>
			<td style="width:275px;">Aldrich</td>
			<td style="width:136px;">115150</td>
			<td style="width:259px;">Note: irritrant to skin; toxic if inhaled</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">Triethylamine</td>
			<td style="width:275px;">Alfa Aesar</td>
			<td style="width:136px;">A12646</td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">Dichloromethane&#160;</td>
			<td style="width:275px;">Acros Organics</td>
			<td style="width:136px;">348460010</td>
			<td style="width:259px;">Extra dry&#160;</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">Magnetic stirrer</td>
			<td style="width:275px;">any source</td>
			<td style="width:136px;"></td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">PAMAM G4 Dendrimer</td>
			<td style="width:275px;">Andrews ChemService</td>
			<td style="width:136px;">AuCS - 297</td>
			<td style="width:259px;">&#160;10 % wt. solution in MeOH</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">Lipophylic Sephadex LH-20</td>
			<td style="width:275px;">Sigma</td>
			<td style="width:136px;">LH20100</td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">Thin-layer chromatography plates</td>
			<td style="width:275px;">Merck Millipore</td>
			<td style="width:136px;">1.05554.0001</td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">Formic acid</td>
			<td style="width:275px;">VWR Chemicals</td>
			<td style="width:136px;">20318.297</td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">Lophylizer&#160;</td>
			<td style="width:275px;">any source</td>
			<td style="width:136px;"></td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:251px;">Gadollinium(III) chloride hexahydrate</td>
			<td style="width:275px;">Aldrich</td>
			<td style="width:136px;">G7532</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">Sodium hydroxide</td>
			<td style="width:275px;">Acros Organics</td>
			<td style="width:136px;">134070010</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">pH meter</td>
			<td style="width:275px;">any source</td>
			<td style="width:136px;"></td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:251px;">Ethylenediaminetetraacetic acid disodium salt dihydrate</td>
			<td style="width:275px;">Aldrich</td>
			<td style="width:136px;">E5134</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">Mass spectrometer (ESI)</td>
			<td style="width:275px;">Agilent</td>
			<td style="width:136px;"></td>
			<td>Ion trap SL 1100&#160;</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">Acetate buffer</td>
			<td style="width:275px;">any source</td>
			<td style="width:136px;"></td>
			<td>pH 5.8</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">Xylenol orange</td>
			<td style="width:275px;">Aldrich</td>
			<td style="width:136px;">52097</td>
			<td>20 &#956;M in acetate buffer</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">Hydrophylic Sephadex G-15</td>
			<td style="width:275px;">GE Healthcare</td>
			<td style="width:136px;">17-0020-01</td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:251px;">Amicon Ultra-15 Centrifugal Filter Unit</td>
			<td style="width:275px;">Merck Millipore</td>
			<td style="width:136px;">UFC900324</td>
			<td style="width:259px;">Ultracel-3 membrane (MWCO 3000)</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">Centrifuge</td>
			<td style="width:275px;">any source</td>
			<td style="width:136px;"></td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">NMR spectrometer&#160;</td>
			<td style="width:275px;">Bruker</td>
			<td style="width:136px;"></td>
			<td style="width:259px;">Avance III 300 MHz</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">Topspin</td>
			<td style="width:275px;">Bruker</td>
			<td style="width:136px;"></td>
			<td style="width:259px;">version 2.1</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">Combustion analysis instrument</td>
			<td style="width:275px;">EuroVector SpA</td>
			<td style="width:136px;"></td>
			<td style="width:259px;">EuroEA 3000 Elemental Analyser&#160;</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">MALDI-ToF MS instrument</td>
			<td style="width:275px;">Applied Biosystems</td>
			<td style="width:136px;"></td>
			<td style="width:259px;">Voyager-STR</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">Deuteriumoxid</td>
			<td style="width:275px;">Carl Roth GmbH</td>
			<td style="width:136px;">6672.3</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">tert-Butyl alcohol</td>
			<td style="width:275px;">Carl Roth GmbH</td>
			<td style="width:136px;">AE16.1</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">Vortex mixer</td>
			<td style="width:275px;">any source</td>
			<td style="width:136px;"></td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">Norell NMR tubes</td>
			<td style="width:275px;">Deutero GmbH</td>
			<td style="width:136px;">507-HP-7</td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">NMR coaxial tube</td>
			<td style="width:275px;">Deutero GmbH</td>
			<td style="width:136px;">coaxialb-5-7</td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">DLS instrument</td>
			<td style="width:275px;">Malvern</td>
			<td style="width:136px;"></td>
			<td style="width:259px;">Zetasizer Nano ZS</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">0.20 &#956;m PTFE filter&#160;</td>
			<td style="width:275px;">Carl Roth GmbH</td>
			<td style="width:136px;">KC94.1</td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">HEPES</td>
			<td style="width:275px;">Fisher BioReagents</td>
			<td style="width:136px;">BP310</td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">Plastic tube vials</td>
			<td style="width:275px;">any source</td>
			<td style="width:136px;"></td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">Dotarem</td>
			<td style="width:275px;">Guerbet</td>
			<td style="width:136px;"></td>
			<td style="width:259px;">NDC 67684-2000-1</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:251px;">MRI scanner</td>
			<td style="width:275px;">Bruker</td>
			<td style="width:136px;"></td>
			<td style="width:259px;">BioSpec 70/30 USR magnet (7 T). Note: potential hazards related to high magnetic fields</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:251px;">RF coil</td>
			<td style="width:275px;">Bruker</td>
			<td style="width:136px;"></td>
			<td style="width:259px;">dual frequency volume coil (RF RES 300 1H/19F 075/040 LIN/LIN TR)</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:251px;">Paravision (software)</td>
			<td style="width:275px;">Bruker</td>
			<td style="width:136px;"></td>
			<td style="width:259px;">Version 5.1</td>
		</tr>
	</tbody>
</table>

preparation and in vitro characterization of dendrimer-based contrast agents for magnetic resonance imaging

비 고리 또는 마크로 킬레이트와 가돌리늄 (III) 상자성의 착물은 자기 공명 영상 (MRI)에 대한 가장 일반적으로 사용되는 조영제 (CAS)이다. 그들의 목적 따라서 MR 화상 콘트라스트 및 MRI 측정의 특이성을 증가 조직 물의 양자의 이완 율을 향상시킬 수있다. 현재 임상 적으로 승인 된 조영제 빠르게 본체로부터 클리어 저 분자량 분자이다. 상자성 킬레이트 제의 담체로서 덴드리머의 사용이 더 효율적인 MRI 조영제의 미래 개발에 중요한 역할을 할 수있다. 특히, 높은 신호 대비 상자성 종 결과의 국소 농도의 증가. 또한,이 CA는 고 분자량과 크기 때문에 장기간 조직 체류 시간을 제공한다. 여기서는 폴리 (아미도 아민)에 기초한 고분자 MRI 조영제 (PAMAM) monomacro 함께 덴드리머를 제조하기위한 편리한 방법을 보여순환 DOTA 형 킬레이트 (DOTA - 1,4,7,10-tetraazacyclododecane-1,4,7,10-테트라). 킬레이트 부 티오 브리지를 형성 PAMAM 덴드리머의 아민 표면기를 향해 이소 티오 시아 네이트 (NCS) 기의 반응성을 이용하여 추가 하였다. 덴드리머 제품 그래피, 핵 자기 공명 스펙트럼, 질량 분석, 및 원소 분석에 의해 분석 하였다. 마지막으로, 고해상도 자기 공명 영상을 기록하고, 제조 된 덴드리머와 시판 제제 단량체로부터 얻어지는 신호 대조와 비교 하였다.

DCA의 제조는 두 단계로 구성되었다 : 1) 단량체 DOTA 형 킬레이트의 합성 (도 1)과 G4의 PAMAM 덴드리머와 복잡 덴드리머 하나님 (III)의 후속 제조와 킬레이트 2) 커플 링 (도 2) . 첫 번째 단계에서, 네 카복실산 함유 cyclen 기반 DOTA 형 킬레이트 및 상기 합성 변형 적합한 직교 그룹을 제조 하였다. 부틸 2- 브로 모 -4- (4- 니트로 페닐) 부?...

Watch this Scientific Journal Video about Preparation and In Vitro Characterization of Dendrimer-based Contrast Agents for Magnetic Resonance Imaging at JoVE.com

Preparation and In Vitro Characterization of Dendrimer-based Contrast Agents for Magnetic Resonance Imaging

이 프로토콜은 상자성 가돌리늄 이온을 배위 cyclen 계 마크로 킬레이트를 운반하는 덴드리머 자기 공명 영상 (MRI) 조영제의 제조 및 특성을 설명한다. 시판되는 모노머 아날로그에 비해 시험 관내 MRI의 일련의 실험에서이 에이전트는 MRI 증폭 신호를 생성했다.

준비 및<em&gt; 체외</em&gt; 자기 공명 영상에 대한 덴드리머 기반 대조 대리인의 특성

Paramagnetic complexes of gadolinium(III) with acyclic or macrocyclic chelates are the most commonly used contrast agents (CAs) for magnetic resonance ...

preparation-vitro-characterization-dendrimer-based-contrast-agents

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JoVE Journal

Chemistry

Preparation and In Vitro Characterization of Dendrimer-based Contrast Agents for Magnetic Resonance Imaging

9.8K 조회수.  Max Planck Institute for Biological Cybernetics.  이 프로토콜은 상자성 가돌리늄 이온을 배위 cyclen 계 마크로 킬레이트를 운반하는 덴드리머 자기 공명 영상 (MRI) 조영제의 제조 및 특성을 설명한다. 시판되는 모노머 아날로그에 비해 시험 관내 MRI의 일련의 실험에서이 에이전트는 MRI 증폭 신호를 생성했다. 

비디오: Preparation and In Vitro Characterization of Dendrimer-based Contrast Agents for Magnetic Resonance Imaging

Synthesis, Cellular Delivery and In vivo Application of Dendrimer-based pH Sensors

The development of fluorescent indicators represented a revolution for life sciences. Genetically encoded and synthetic fluorophores with sensing abilities allowed the visualization of biologically relevant species with high spatial and temporal resolution. Synthetic dyes are of particular interest thanks to their high tunability and the wide range of measureable analytes. However, these molecules suffer several limitations related to small molecule behavior (poor solubility, difficulties in targeting, often no ratiometric imaging allowed). In this work we introduce the development of dendrimer-based sensors and present a procedure for pH measurement in vitro, in living cells and in vivo. We choose dendrimers as ideal platform for our sensors for their many desirable properties (monodispersity, tunable properties, multivalency) that made them a widely used scaffold for several biomedical devices. The conjugation of fluorescent pH indicators to the dendrimer scaffold led to an enhancement of their sensing performances. In particular dendrimers exhibit reduced cell leakage, improved intracellular targeting and allow ratiometric measurements. These novel sensors were successfully employed to measure pH in living HeLa cells and in vivo in mouse brain.

Synthesis, Cellular Delivery and In vivo Application of Dendrimer-based pH Sensors

Fluorescence sensors are powerful tools in life science. Here we describe a methodology to synthesize and use dendrimer-based fluorescent sensors to measure pH in living cells and in vivo. The dendritic scaffold enhances the properties of conjugated fluorescent dyes leading to improved sensing properties.

합성, 세포 전달 및<em&gt; 생체 내</em&gt; 덴드리머 기반 pH 센서의 응용 프로그램

The development of  fluorescent indicators represented a revolution for life sciences. Genetically  encoded and synthetic fluorophores with sensing ...

연구

화학

Preparation and Characterization of SDF-1&#945;-Chitosan-Dextran Sulfate Nanoparticles

Chitosan (CS) and dextran sulfate (DS) are charged polysaccharides (glycans), which form polyelectrolyte complex-based nanoparticles when mixed under appropriate conditions. The glycan nanoparticles are useful carriers for protein factors, which facilitate the in vivo delivery of the proteins and sustain their retention in the targeted tissue. The glycan polyelectrolyte complexes are also ideal for protein delivery, as the incorporation is carried out in aqueous solution, which reduces the likelihood of inactivation of the proteins. Proteins with a heparin-binding site adhere to dextran sulfate readily, and are, in turn, stabilized by the binding. These particles are also less inflammatory and toxic when delivered in vivo. In the protocol described below, SDF-1&#945; (Stromal cell-derived factor-1&#945;), a stem cell homing factor, is first mixed and incubated with dextran sulfate. Chitosan is added to the mixture to form polyelectrolyte complexes, followed by zinc sulfate to stabilize the complexes with zinc bridges. The resultant SDF-1&#945;-DS-CS particles are measured for size (diameter) and surface charge (zeta potential). The amount of the incorporated SDF-1&#945; is determined, followed by measurements of its in vitro release rate and its chemotactic activity in a particle-bound form.

The objective of this protocol is to incorporate SDF-1&#945;, a stem cell homing factor, into dextran sulfate-chitosan nanoparticles. The resultant particles are measured for their size and zeta potential, as well as the content, activity, and in vitro release rate of SDF-1&#945; from the nanoparticles.

SDF-1α-키토산 덱스 트란 황산 나노 입자의 제조 및 특성

Chitosan (CS) and dextran sulfate (DS) are charged polysaccharides (glycans), which form polyelectrolyte complex-based nanoparticles when mixed under ...

Preparation and Characterization of Individual and Multi-drug Loaded Physically Entrapped Polymeric Micelles

Amphiphilic block copolymers like polyethyleneglycol-block-polylactic acid (PEG-b-PLA) can self-assemble into micelles above their critical micellar concentration forming hydrophobic cores surrounded by hydrophilic shells in aqueous environments. The core of these micelles can be utilized to load hydrophobic, poorly water soluble drugs like docetaxel (DTX) and everolimus (EVR). Systematic characterization of the micelle structure and drug loading capabilities are important before in vitro and in vivo studies can be conducted. The goal of the protocol described herein is to provide the necessary characterization steps to achieve standardized micellar products. DTX and EVR have intrinsic solubilities of 1.9 and 9.6 &#181;g/ml respectively Preparation of these micelles can be achieved through solvent casting which increases the aqueous solubility of DTX and EVR to 1.86 and 1.85 mg/ml, respectively. Drug stability in micelles evaluated at room temperature over 48 hr indicates that 97% or more of the drugs are retained in solution. Micelle size was assessed using dynamic light scattering and indicated that the size of these micelles was below 50 nm and depended on the molecular weight of the polymer. Drug release from the micelles was assessed using dialysis under sink conditions at pH 7.4 at 37 oC over 48 hr. Curve fitting results indicate that drug release is driven by a first order process indicating that it is diffusion driven.

The goal of this protocol is to describe the preparation and characterization of physically entrapped, poorly water soluble drugs in micellar drug delivery systems composed of amphiphilic block copolymers.

준비 및 개인의 특성과 다중 약물은 물리적으로 고분자 미셀을 포획로드

Amphiphilic block copolymers like polyethyleneglycol-block-polylactic acid (PEG-b-PLA) can self-assemble into micelles above their critical micellar ...

Sequencing of Plant Wall Heteroxylans Using Enzymic, Chemical (Methylation) and Physical (Mass Spectrometry, Nuclear Magnetic Resonance) Techniques

This protocol describes the specific techniques used for the characterization of reducing end (RE) and internal region glycosyl sequence(s) of heteroxylans. De-starched wheat endosperm cell walls were isolated as an alcohol-insoluble residue (AIR)1 and sequentially extracted with water (W-sol Fr) and 1 M KOH containing 1% NaBH4 (KOH-sol Fr) as described by Ratnayake et al. (2014)2. Two different approaches (see summary in Figure 1) are adopted. In the first, intact W-sol AXs are treated with 2AB to tag the original RE backbone chain sugar residue and then treated with an endoxylanase to generate a mixture of 2AB-labelled RE and internal region reducing oligosaccharides, respectively. In a second approach, the KOH-sol Fr is hydrolyzed with endoxylanase to first generate a mixture of oligosaccharides which are subsequently labelled with 2AB. The enzymically released ((un)tagged) oligosaccharides from both W- and KOH-sol Frs are then methylated and the detailed structural analysis of both the native and methylated oligosaccharides is performed using a combination of MALDI-TOF-MS, RP-HPLC-ESI-QTOF-MS and ESI-MSn. Endoxylanase digested KOH-sol AXs are also characterized by nuclear magnetic resonance (NMR) that also provides information on the anomeric configuration. These techniques can be applied to other classes of polysaccharides using the appropriate endo-hydrolases.

Sequencing of Plant Wall Heteroxylans Using Enzymic, Chemical Methylation and Physical Mass Spectrometry, Nuclear Magnetic Resonance Techniques

This protocol describes the specific techniques used for the structural characterization of reducing end (RE) and internal region glycosyl sequence(s) of heteroxylans by tagging the RE with 2 aminobenzamide prior to enzymatic (endoxylanase) hydrolysis and then analysis of the resultant oligosaccharides using mass spectrometry (MS) and nuclear magnetic resonance (NMR).

This protocol describes the specific techniques used for the characterization of reducing end (RE) and internal region glycosyl sequence(s) of ...

Synthesis and Characterization of Supramolecular Colloids

Control over colloidal assembly is of utmost importance for the development of functional colloidal materials with tailored structural and mechanical properties for applications in photonics, drug delivery and coating technology. Here we present a new family of colloidal building blocks, coined supramolecular colloids, whose self-assembly is controlled through surface-functionalization with a benzene-1,3,5-tricarboxamide (BTA) derived supramolecular moiety. Such BTAs interact via directional, strong, yet reversible hydrogen-bonds with other identical BTAs. Herein, a protocol is presented that describes how to couple these BTAs to colloids and how to quantify the number of coupling sites, which determines the multivalency of the supramolecular colloids. Light scattering measurements show that the refractive index of the colloids is almost matched with that of the solvent, which strongly reduces the van der Waals forces between the colloids. Before photo-activation, the colloids remain well dispersed, as the BTAs are equipped with a photo-labile group that blocks the formation of hydrogen-bonds. Controlled deprotection with UV-light activates the short-range hydrogen-bonds between the BTAs, which triggers the colloidal self-assembly. The evolution from the dispersed state to the clustered state is monitored by confocal microscopy. These results are further quantified by image analysis with simple routines using ImageJ and Matlab. This merger of supramolecular chemistry and colloidal science offers a direct route towards light- and thermo-responsive colloidal assembly encoded in the surface-grafted monolayer.

A protocol for the synthesis and characterization of colloids coated with supramolecular moieties is described. These supramolecular colloids undergo self-assembly upon the activation of the hydrogen-bonds between the surface-anchored molecules by UV-light.

Control over colloidal assembly is of utmost importance for the development of functional colloidal materials with tailored structural and mechanical ...

Low-energy Cathodoluminescence for (Oxy)Nitride Phosphors

Nitride and oxynitride (Sialon) phosphors are good candidates for the ultraviolet and visible emission applications. High performance, good stability and flexibility of their emission properties can be achieved by controlling their composition and dopants. However, a lot of work is still required to improve their properties and to reduce the production cost. A possible approach is to correlate the luminescence properties of the Sialon particles with their local structural and chemical environment in order to optimize their growth parameters and find novel phosphors. For such a purpose, the low-voltage cathodoluminescence (CL) microscopy is a powerful technique. The use of electron as an excitation source allows detecting most of the luminescence centers, revealing their luminescence distribution spatially and in depth, directly comparing CL results with the other electron-based techniques, and investigating the stability of their luminescence properties under stress. Such advantages for phosphors characterization will be highlighted through examples of investigation on several Sialon phosphors by low-energy CL.

Low-energy Cathodoluminescence for OxyNitride Phosphors

An excellent chemical and luminescence stabilities of (oxy)nitride phosphors present it as an promising alternative to currently used sulfide and oxide phosphors. In this paper, we present the way to investigate its local luminescence properties using low-energy cathodoluminescence (CL).

(옥시) 질화물 형광체에 대한 낮은 에너지 음극선

Nitride and oxynitride (Sialon) phosphors are good candidates for the ultraviolet and visible emission applications. High performance, good stability and ...

In Situ Monitoring of Diffusion of Guest Molecules in Porous Media Using Electron Paramagnetic Resonance Imaging

A method is demonstrated to monitor macroscopic translational diffusion using electron paramagnetic resonance (EPR) imaging. A host-guest system with nitroxide spin probe 3-(2-Iodoacetamido)-2,2,5,5-tetramethyl-1-pyrrolidinyloxy (IPSL) as a guest inside the periodic mesoporous organosilica (PMO) aerogel UKON1-GEL as a host and ethanol as a solvent is used as an example to describe the protocol. Data is shown from a previous publication, where the protocol has been applied to both IPSL and Tris(8-carboxy-2,2,6,6-perdeutero-tetramethyl-benzo[1,2-d:4,5-d&#8242;]bis(1,3)dithiole) methyl (Trityl) as guest molecules and UKON1-GEL and SILICA-GEL as host systems. 
A method is shown to prepare aerogel samples that cannot be synthesized directly in the sample tube for measurement due to a size change during synthesis. The aerogel is attached to sample tubes using heat shrink tubing and a pressure cooker to reach the necessary temperature without evaporating the solvent in the process. The method does not assume a clearly defined initial distribution of guest molecules at the start of the measurement. Instead, it requires a reservoir on top of the aerogel and experimentally determines the influx rate during data analysis.
The diffusion is monitored continually over a period of 20 hr by recording the 1d spin density profile within the sample. The spectrometer settings for the imaging experiment are described quantitatively. Data analysis software is provided to take the resonator sensitivity profile into account and to numerically solve the diffusion equation. The software determines the macroscopic translational diffusion coefficient by least square minimization of the difference between the experiment and the numerical solution of the diffusion equation.

In Situ Monitoring of Diffusion of Guest Molecules in Porous Media Using Electron Paramagnetic Resonance Imaging

A protocol for the in situ monitoring of the diffusion of guest molecules in porous media using electron paramagnetic resonance (EPR) imaging is presented.

전자 상자성 공명 영상을 이용하여 다공성 미디어에서 고객 분자의 확산의 제자리 모니터링에

A method is demonstrated to monitor macroscopic translational diffusion using electron paramagnetic resonance (EPR) imaging. A host-guest system with ...

DNA-magnetic Particle Binding Analysis by Dynamic and Electrophoretic Light Scattering

Isolation of DNA using magnetic particles is a field of high importance in biotechnology and molecular biology research. This protocol describes the evaluation of DNA-magnetic particles binding via dynamic light scattering (DLS) and electrophoretic light scattering (ELS). Analysis by DLS provides valuable information on the physicochemical properties of particles including particle size, polydispersity, and zeta potential. The latter describes the surface charge of the particle which plays major role in electrostatic binding of materials such as DNA. Here, a comparative analysis exploits three chemical modifications of nanoparticles and microparticles and their effects on DNA binding and elution. Chemical modifications by branched polyethylenimine, tetraethyl orthosilicate and (3-aminopropyl)triethoxysilane are investigated. Since DNA exhibits a negative charge, it is expected that zeta potential of particle surface will decrease upon binding of DNA. Forming of clusters should also affect particle size. In order to investigate the efficiency of these particles in isolation and elution of DNA, the particles are mixed with DNA in low pH (~6), high ionic strength and dehydration environment. Particles are washed on magnet and then DNA is eluted by Tris-HCl buffer (pH = 8). DNA copy number is estimated using quantitative polymerase chain reaction (PCR). Zeta potential, particle size, polydispersity and quantitative PCR data are evaluated and compared. DLS is an insightful and supporting method of analysis that adds a new perspective to the process of screening of particles for DNA isolation.

This protocol describes the synthesis of magnetic particles and evaluation of their DNA-binding properties via dynamic and electrophoretic light scattering. This method focuses on monitoring changes in particle size, their polydispersity and zeta potential of particle surface which play major role in binding of materials such as DNA.

역동적이 고 전기 이동 산란 분석 바인딩 DNA-마그네틱 입자

Isolation of DNA using magnetic particles is a field of high importance in biotechnology and molecular biology research. This protocol describes the ...

Preparation and Characterization of C60/Graphene Hybrid Nanostructures

Physical thermal deposition in a high vacuum environment is a clean and controllable method for fabricating novel molecular nanostructures on graphene. We present methods for depositing and passively manipulating C60 molecules on rippled graphene that advance the pursuit of realizing applications involving 1D C60/graphene hybrid structures. The techniques applied in this exposition are geared towards high vacuum systems with preparation areas capable of supporting molecular deposition as well as thermal annealing of the samples. We focus on C60 deposition at low pressure using a homemade Knudsen cell connected to a scanning tunneling microscopy (STM) system. The number of molecules deposited is regulated by controlling the temperature of the Knudsen cell and the deposition time. One-dimensional (1D) C60 chain structures with widths of two to three molecules can be prepared via tuning of the experimental conditions. The surface mobility of the C60 molecules increases with annealing temperature allowing them to move within the periodic potential of the rippled graphene. Using this mechanism, it is possible to control the transition of 1D C60 chain structures to a hexagonal close packed quasi-1D stripe structure.

Preparation and Characterization of C60/Graphene Hybrid Nanostructures

Here we present a protocol for the fabrication of C60/graphene hybrid nanostructures by physical thermal evaporation. Particularly, the proper manipulation of deposition and annealing conditions allow the control over the creation of 1D and quasi 1D C60 structures on rippled graphene.

준비 및 C60/Graphene 하이브리드 Nanostructures의 특성화

Physical thermal deposition in a high vacuum environment is a clean and controllable method for fabricating novel molecular nanostructures on graphene. We ...

Photoelectron Imaging of Anions Illustrated by 310 Nm Detachment of F&#8722;

Anion photoelectron imaging is a very efficient method for the study of energy states of bound negative ions, neutral species and interactions of unbound electrons with neutral molecules/atoms. State-of-the-art in vacuo anion generation techniques allow application to a broad range of atomic, molecular, and cluster anion systems. These are separated and selected using time-of-flight mass spectrometry. Electrons are removed by linearly polarized photons (photo detachment) using table-top laser sources which provide ready access to excitation energies from the infra-red to the near ultraviolet. Detecting the photoelectrons with a velocity mapped imaging lens and position sensitive detector means that, in principle, every photoelectron reaches the detector and the detection efficiency is uniform for all kinetic energies. Photoelectron spectra extracted from the images via mathematical reconstruction using an inverse Abel transformation reveal details of the anion internal energy state distribution and the resultant neutral energy states. At low electron kinetic energy, typical resolution is sufficient to reveal energy level differences on the order of a few millielectron-volts, i.e., different vibrational levels for molecular species or spin-orbit splitting in atoms. Photoelectron angular distributions extracted from the inverse Abel transformation represent the signatures of the bound electron orbital, allowing more detailed probing of electronic structure. The spectra and angular distributions also encode details of the interactions between the outgoing electron and the residual neutral species subsequent to excitation. The technique is illustrated by the application to an atomic anion (F&#8722;), but it can also be applied to the measurement of molecular anion spectroscopy, the study of low lying anion resonances (as an alternative to scattering experiments) and femtosecond (fs) time resolved studies of the dynamic evolution of anions.

Here, we present a protocol for photoelectron imaging of anionic species. Anions generated in vacuo and separated by mass spectrometry are probed using velocity mapped photoelectron imaging, providing details of anion and neutral energy levels, anion and neutral structure and the nature of the anion electronic state.

F− 의 310 Nm 분리에 의해 설명 하는 음이온의 광전자 이미징

Anion photoelectron imaging is a very efficient method for the study of energy states of bound negative ions, neutral species and interactions of unbound ...