我々は彼の親切なサポートのために教授JPスパッツ（生物物理化学、ハイデルベルク大学、新素材やバイオシステムズの専攻、知能システムのためのマックスプランク研究所、シュツットガルト）に感謝。マックスプランク研究機構とドイツ学術振興（EAC-AへのDFG SFB/TR79。）からの財政支援も大幅に認められている。

1。 11-Mercaptoundecanoyl-N-ヒドロキシスクシンイミドエステル（MU-NHS）の合成<ol><li>室温（RT）で40 mlのジクロロメタン（PA）中の10 mlのアセトン（PA）（ジメチルアミノ）ピリジン1gの11 -メルカプトウンデカン酸- 500mgのN-ヒドロキシスクシンイミドの溶液及び30mgの4滴ずつ加える。 </li><li> 0℃に反応液を冷却し、（窒素雰囲気下）ジクロロメタン10mlを滴下1.1グラムN、N &#39; -ジシクロヘキシルカルボジイミドを加える。 1時間低温で反応を維持し、次いで室温で一晩撹拌する。 </li><li>沈殿物を濾過し、減圧下で乾燥させます。 1:1の割合で石油ベンゼン、酢酸エチル（PA）を用いるフラッシュクロマトグラフィーによって生成物を精製する。 </li></ol> 2。均質な金層の作製<ol><li>精密ワイプとし、酢酸エチル（PA）およびメタノール（PA）の1:1混合物を含有する溶液中で5分間超音波処理して清浄なガラスカバースリップ。 RinsE基質メタノールで、窒素気流下で乾燥させます。 </li><li>スパッタコーティング装置ではクリーンな基板を配置します。チャンバを排気。 60秒間スパッタ法により、クロムターゲットから最上部のクロム酸化物層を除去。金属線と40 nmの金の層を塗布し、その後10nmのクロム層でコートして下にサンプルを置きます。 </li></ol> 3。 BMP-2の表面固定化<ol><li>約1mMの最終濃度までNにおけるMU-NHS、N-dimethlyformamide（DMF）に溶解し、窒素雰囲気下で4時間RTにてMU-NHS溶液中に金基板をインキュベートする。 </li><li> 2分間DMF中で超音波処理表面は、DMF、MeOHで洗浄し、窒素気流下で乾燥させます。 </li><li> 100μg/ mlの（-80℃で保存アリコート）の濃度で滅菌した4 mMのHCl中のrhBMP-2のストック溶液を準備します。ワーキング希釈液（3.5μg/ ml）をするために、PBS / NaClを（PBSで1 M NaClを含む）と、中のストック溶液を希釈使用直前にpH8.5にdjust。 </li><li>一晩4℃でのrhBMP-2作業溶液中にMU-NHS官能表面をインキュベートする。インキュベーション上清を取り除きます。超音波処理は、2分間、PBS / NaCl中に表面および滅​​菌PBS / NaClで3回洗う。 </li></ol> 4。表面特性化<ol><li>抗体の非特異的吸着をブロックするために、抗BMP-2抗体（1:100、1％（重量/体積とのインキュベーション、続いてRTで1時間PBS溶液中の5％BSA（w / v）ので表面をインキュベートするRTで1時間）BSA / PBS溶液）。 PBSおよび30秒間の超音波処理で二回表面を洗浄します。 </li><li>次いで、PBSで二回洗浄し、RTで30分間HRP結合二次抗体（1:1,000、1％（w / v）のBSA / PBS溶液）で基質をインキュベートする。 </li><li>プレートリーダーを用いて570nmでのHRP酵素活性を測定するためにAmplifluレッドアッセイを使用する。 </li></ol> 5。生物学的活性の解析<ol><li>シード1×10 6ウェルPLにおけるウェルあたり5マウスC2C12筋芽細胞10％FBS、1％ペニシリン/ストレプトマイシンおよび37℃/ 5％CO 2で24時間、それらをインキュベートを補充したピルビン酸塩を含有する高グルコースDMEMからなる増殖培地中でのATE。 </li><li>固定化されたBMP-2で飾られた表面に播種する前3-5時間無血清DMEMで細胞を餓死。 </li><li>短期的なシグナル誘導の調査のために、200μlの新鮮な無血清DMEMで培地を交換し、細胞上に固定化BMP-2の表面に配置します。表面は傷を避けるために、先端の細いピンセットで処理する必要があります。 </li><li>優しく表面を削除し、ピペットで培地を吸引、PBSで2回細胞を洗浄。細胞応答の分析に進みます。 </li><li>長期の生物学的活性の分析のため、細胞をプレート低血清条件（2％FBS）で表面及び37℃/ 5％CO 2で培養して上に6日間。 </li></ol>

<ol>
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著者らは、開示することは何もありません。

このプロトコルでは、生物活性のrhBMP-2で官能化表面の調製が記載されている。 ;のrhBMP-2タンパク質の2）の共有結合固定金表面上の二官能性リンカーの自己集合単分子層（SAM）の1）初期形成：このアプローチは、2つのステップを含む。以前の研究では、二官能性リンカーおよび成長因子の有効な結合を検証し、表面固定化のrhBMP-2は、その生物学的活性19を維持することを実?...

骨形成タンパク質2（BMP-2）は、 デノボ骨形成ならびに胚発生および成体のホメオスタシス1-3中のいくつかの組織の調節因子の誘導物質としてトランスフォーミング成長因子（TGF-β）ファミリーの作用のメンバーである。 生物学的に活性なホモ二量体のBMP-2タンパク質の各単量体は、非常にすべてのBMP 4に保存されている「システインノット」モチーフを、含まれています。第七のシステインは二つのモノマー5,6間の分子間結合を形成する、二量体化に関与しているのに対し、7個のシステイン残基の六つは、各単量体を安定化する分子内ジスルフィド結合を形成する。この高度に保存されたシステインノットは、BMP-2タンパク質の三次元構造を定義し、熱、変性剤及び酸性pH 7-9に対する抵抗としてのそのユニークな特性を決定する。 BMP-2は、それによってシグナル伝達を誘導する、セリン/スレオニンキナーゼ膜貫通受容体に結合する<sup&gt; 10月12日。受容体のオリゴマーのモードに応じて、異なるシグナル伝達経路が活性化されています。のSmad非依存のシグナル伝達カスケードは、Smadの複雑な核移行との転写の活性化受容体リン酸化の結果によって活性化されたSmad依存性経路のに対し、p38シグナルを経由して、アルカリホスファターゼ誘導を導くこのような分化の阻害剤（Id）の12〜14などの特定の標的遺伝子。 骨形態では、BMP-2は、このように治癒および骨のデノボ形成を刺激、骨芽細胞への間葉系幹細胞の分化を誘導する。現在、組換え的にBMP-2は、骨折したサイトの治癒を促進するために臨床応用されている表現。骨組織工学における一般的な方法は、局所送達システムと比較して低侵襲性で注射可能な成長因子の使用である。しかしながら、 インビボ研究と臨床応用が示されているその短い生物学的半減期、非特異的locのalizationおよびBMP-2の迅速な局所的クリアランスは、いくつかの、地元の異所性および全身の問題15につながる可能性があります。従って、効果的なプレゼンテーションを得るために、材料内又は上にBMP-2の捕捉または固定化は、標的部位でのそのローカルおよび持続送達のために必要である。持続的な送達は、物理的捕捉、吸着またはイオン錯体16のような非共有結合性の保持の手法を用いて達成することができる。しかしながら、分子17の変性における表面へのタンパク質の非特異的吸着がよい結果が知られている。増殖因子の共有結合のために、支持体の種類は、過去10年間にわたって開発されてきた。例えば、タンパク質のアミノ基またはカルボキシル基を標的とする二官能性連結分子の使用は、必ずしも固定化を達成するために、タンパク質の修飾を必要としないアプローチの一種である。実際には、一方のタンパク質修飾は、タンパク質の配向を制御するという利点を提供人工的なドメイン、ペプチドタグおよび部位特異的鎖の導入は、成長の生物学的活性は、17因子変更することができる。従って、原因支持材料との相互作用のために変性を回避するために、表面が所望の因子18のカップリングに続いて、連結分子の自己組織化単分子膜（SAM）を用いて、例えば、予め官能化することができる。我々は、その遊離アミン残基を標的とすることによって共有結合的に表面にBMP-2を固定化するSAMベースのアプローチを使用して、固定化タンパク質がその短期および長期の生物学的活性の19の両方を保持することが示されている。このプロトコルは、細胞膜で発生し、骨形成シグナルを担当する細胞内シグナル伝達を調節するメカニズムに関するin vitro試験のための細胞にBMP-2を実現するための簡単で効率的な方法を提供します。

<table border="1">
	<tbody>
		<tr>
			<td>Name of Material/ Equipment</td>
			<td>Company</td>
			<td>Catalog Number</td>
			<td>Comments/Description</td>
		</tr>
		<tr>
			<td>N-hydroxysuccinimide</td>
			<td>Sigma-Aldrich</td>
			<td>130672</td>
			<td></td>
		</tr>
		<tr>
			<td>4-(dimethylamino)pyridin</td>
			<td>Sigma-Aldrich</td>
			<td>522805</td>
			<td></td>
		</tr>
		<tr>
			<td>Acetone</td>
			<td>AppliChem</td>
			<td>A2282</td>
			<td></td>
		</tr>
		<tr>
			<td>11-mercaptoundecanoic acid</td>
			<td>Sigma-Aldrich</td>
			<td>674427</td>
			<td></td>
		</tr>
		<tr>
			<td>Dichlormethane</td>
			<td>Merck</td>
			<td>106050</td>
			<td></td>
		</tr>
		<tr>
			<td>N,N&#39;-dicyclohexylcarbodiimide</td>
			<td>Sigma-Aldrich</td>
			<td>D80002</td>
			<td></td>
		</tr>
		<tr>
			<td>Petroleum benzene</td>
			<td>Merck</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Glass coverslips</td>
			<td>Carl Roth</td>
			<td>M 875</td>
			<td></td>
		</tr>
		<tr>
			<td>Ethylacetate</td>
			<td>AppliChem</td>
			<td>A3550</td>
			<td></td>
		</tr>
		<tr>
			<td>Methanol</td>
			<td>Carl Roth</td>
			<td>4627</td>
			<td></td>
		</tr>
		<tr>
			<td>N,N-dimethylformamide</td>
			<td>Carl Roth</td>
			<td>T921</td>
			<td></td>
		</tr>
		<tr>
			<td>rhBMP-2</td>
			<td>R&#38;D Systems</td>
			<td>355-BM</td>
			<td>Carrier-free; expressed in E.coli</td>
		</tr>
		<tr>
			<td>PBS</td>
			<td>PAA</td>
			<td>H15-002</td>
			<td></td>
		</tr>
		<tr>
			<td>NaCl</td>
			<td>Carl Roth</td>
			<td>HN00.2</td>
			<td></td>
		</tr>
		<tr>
			<td>Poly(dimethyl siloxane) (PDMS)</td>
			<td>Dow Corning</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Sylgard 184 silicone elastomer kit</td>
			<td>Dow Corning</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Anti-rhBMP-2</td>
			<td>Sigma</td>
			<td>B9553</td>
			<td></td>
		</tr>
		<tr>
			<td>Goat anti-mouse IgG-HRP</td>
			<td>Santa Cruz</td>
			<td>sc-2005</td>
			<td>Secondary antibody</td>
		</tr>
		<tr>
			<td>Ampliflu Red assay</td>
			<td>Sigma</td>
			<td>90101</td>
			<td></td>
		</tr>
		<tr>
			<td>Dulbecco&#39;s Modified Eagle Medium (DMEM) (1x), liquid</td>
			<td>Gibco</td>
			<td>41966</td>
			<td>High glucose</td>
		</tr>
		<tr>
			<td>Fetal Bovine Serum (FBS)</td>
			<td>Sigma</td>
			<td>F7524</td>
			<td>Sterile filtered, cell culture tested</td>
		</tr>
		<tr>
			<td>Pen/Strep</td>
			<td>Gibco</td>
			<td>15140</td>
			<td></td>
		</tr>
		<tr>
			<td>Trypsin 0.05% (1x) with EDTA 4Na</td>
			<td>Gibco</td>
			<td>25300</td>
			<td></td>
		</tr>
		<tr>
			<td>Glycine (0.1 M)</td>
			<td>Riedel-de Ha&#235;n</td>
			<td>33226</td>
			<td></td>
		</tr>
		<tr>
			<td>IGEPAL CA-630 (1%)</td>
			<td>Sigma</td>
			<td>I8896</td>
			<td>Lysis buffer (ALP assay)19</td>
		</tr>
		<tr>
			<td>Magnesium chloride (MgCl2)(1 mM)</td>
			<td>Carl Roth</td>
			<td>HNO3.2</td>
			<td></td>
		</tr>
		<tr>
			<td>Zinc chloride (ZnCl2) (1 mM)</td>
			<td>Carl Roth</td>
			<td>3533.1</td>
			<td></td>
		</tr>
		<tr>
			<td>p-nitrophenylphosphate (pNPP)</td>
			<td>Sigma</td>
			<td>S0942</td>
			<td>Phosphatase substrate</td>
		</tr>
		<tr>
			<td>Anti-mysin heavy chain (MHC)</td>
			<td>Developmental Studies Hybridoma Bank, University of Iowa</td>
			<td>MF20</td>
			<td>Monoclonal antibody</td>
		</tr>
		<tr>
			<td>Alexa Fluor 488 Goat anti-mouse IgG</td>
			<td>Invitrogen</td>
			<td>A11001</td>
			<td></td>
		</tr>
		<tr>
			<td>DAPI</td>
			<td>Sigma</td>
			<td>D9542</td>
			<td></td>
		</tr>
		<tr>
			<td></td>
			<td></td>
			<td></td>
			<td>Equipment</td>
		</tr>
		<tr>
			<td>Ultrsonic bath (Sonorex Super RK 102H), Frequency 35 kHz</td>
			<td>BANDELIN electronic GmbH &#38; Co. KG</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>MED 020 Sputtercoating system</td>
			<td>BAL-TEC AG</td>
			<td></td>
			<td>Coating conditions 
			Cr: 120 mA, 1.3 x 10-2 mbar, 30 sec 
			Au: 60 mA, 5.0 x 10-2 mbar, 45 sec</td>
		</tr>
		<tr>
			<td>Tecan Infinite M200 Plate reader</td>
			<td>Tecan</td>
			<td></td>
			<td></td>
		</tr>
	</tbody>
</table>

covalent binding of bmp-2 on surfaces using a self-assembled monolayer approach

骨形成タンパク質2（BMP-2）は、骨組織の細胞外マトリックス中に埋め込まれた成長因子である。このようにして治癒およびデノボ骨形成を刺激する骨芽細胞への間葉系細胞の分化のトリガーとしてBMP-2作用する。足場と組み合わせた組換えヒトBMP-2（のrhBMP-2）の臨床使用は、プレゼンテーションモードに基づいて、最近の論争を調達していると量が送達される。ここで紹介するプロトコルは、細胞へのin vitroでの研究のために、BMP-2を実現するための簡単で効率的な方法を提供します。私たちは、ヘテロ二官能性リンカーからなる自己組織化単分子膜を形成する方法を説明し、のrhBMP-2の共有結合固定化を得るために、その後の結合工程を示す。このアプローチでは、タンパク質の生物学的活性を維持しながら、BMP-2の持続的な提示を実現することができる。実際には、BMP-2の表面固定化は、非特異的な広告を防ぐことによって、ターゲットを絞った調査を可能にするorption、成長因子の量を低減しつつ、最も顕著には、表面からの制御されない放出を阻害する。細胞は、共有結合で固定化したrhBMP-2を提示する表面に露出しているときにBMP-2によって引き起こさ両方短期および長期のシグナル伝達事象は、BMP-2刺激に対する細胞応答に関するin vitro研究のためのこのアプローチが適して、行われている。

それは生物学的に不特定の化学的な調整可能なシステムを提供していますので、我々のセットアップでは、金を賦形剤として選ばれた。さらに、自己組織化単分子膜の適用は、多くの利点を伴う。それらの官能末端基をさらに修正することができながら、SAMは自然に、金属や欠陥の少ないフォーム単層上での「ヘッドグループ」を経由して吸着する。こうして彼らは、制御された、まだ適応?...

Watch this Scientific Journal Video about Covalent Binding of BMP-2 on Surfaces Using a Self-assembled Monolayer Approach at JoVE.com

Covalent Binding of BMP-2 on Surfaces Using a Self-assembled Monolayer Approach

我々は、表面上にBMP-2の効率的な固定化を達成するための方法を記載している。我々のアプローチは、その遊離アミン残基を介してBMP-2の共有結合を達成するために、自己組織化単分子膜の形成に基づく。この方法は、細胞膜でのシグナル伝達を研究するための便利なツールです。

共有結合は、自己組織化単分子膜の手法を用いて表面上のBMP-2の結合

Bone morphogenetic protein 2 (BMP-2) is a growth factor embedded in the extracellular matrix of bone tissue. BMP-2 acts as trigger of mesenchymal cell ...

covalent-binding-bmp-2-on-surfaces-using-self-assembled-monolayer

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Unit 1

Covalent Bonding and Structure

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13.8K ビュー.  University of Heidelberg. 我々は、表面上にBMP-2の効率的な固定化を達成するための方法を記載している。我々のアプローチは、その遊離アミン残基を介してBMP-2の共有結合を達成するために、自己組織化単分子膜の形成に基づく。この方法は、細胞膜でのシグナル伝達を研究するための便利なツールです。 

ビデオ: Covalent Binding of BMP-2 on Surfaces Using a Self-assembled Monolayer Approach

Stabilizing Hepatocellular Phenotype Using Optimized Synthetic Surfaces

Currently, one of the major limitations in cell biology is maintaining differentiated cell phenotype. Biological matrices are commonly used for culturing and maintaining primary and pluripotent stem cell derived hepatocytes. While biological matrices are useful, they permit short term culture of hepatocytes, limiting their widespread application. We have attempted to overcome the limitations using a synthetic polymer coating. Polymers represent one of the broadest classes of biomaterials and possess a wide range of mechanical, physical and chemical properties, which can be fine-tuned for purpose. Importantly, such materials can be scaled to quality assured standards and display batch-to-batch consistency. This is essential if cells are to be expanded for high through-put screening in the pharmaceutical testing industry or for cellular based therapy. Polyurethanes (PUs) are one group of materials that have shown promise in cell culture. Our recent progress in optimizing a polyurethane coated surface, for long-term culture of human hepatocytes displaying stable phenotype, is presented and discussed.

This article will focus on developing polymer coated surfaces for long-term, stable culture of stem cell derived human hepatocytes.

最適化された合成表面を使用した肝細胞表現型の安定化

Currently, one of the major limitations in cell biology is maintaining differentiated cell phenotype. Biological matrices are commonly used for culturing ...

化学

An Inverse Analysis Approach to the Characterization of Chemical Transport in Paints

The ability to directly characterize chemical transport and interactions that occur within a material (i.e., subsurface dynamics) is a vital component in understanding contaminant mass transport and the ability to decontaminate materials. If a material is contaminated, over time, the transport of highly toxic chemicals (such as chemical warfare agent species) out of the material can result in vapor exposure or transfer to the skin, which can result in percutaneous exposure to personnel who interact with the material. Due to the high toxicity of chemical warfare agents, the release of trace chemical quantities is of significant concern. Mapping subsurface concentration distribution and transport characteristics of absorbed agents enables exposure hazards to be assessed in untested conditions. Furthermore, these tools can be used to characterize subsurface reaction dynamics to ultimately design improved decontaminants or decontamination procedures. To achieve this goal, an inverse analysis mass transport modeling approach was developed that utilizes time-resolved mass spectroscopy measurements of vapor emission from contaminated paint coatings as the input parameter for calculation of subsurface concentration profiles. Details are provided on sample preparation, including contaminant and material handling, the application of mass spectrometry for the measurement of emitted contaminant vapor, and the implementation of inverse analysis using a physics-based diffusion model to determine transport properties of live chemical warfare agents including distilled mustard (HD) and the nerve agent VX.

In this paper, a procedure for quantifying the mass transport parameters of chemicals in various materials is presented. This process involves employing an inverse-analysis based diffusion model to vapor emission profiles recorded by real-time, mass spectrometry in high vacuum.

塗料中の化学輸送のキャラに逆解析アプローチ

The ability to directly characterize chemical transport and interactions that occur within a material (i.e., subsurface dynamics) is a vital component in ...

Facile Preparation of Internally Self-assembled Lipid Particles Stabilized by Carbon Nanotubes

We present a facile method to prepare nanostructured lipid particles stabilized by carbon nanotubes (CNTs). Single-walled (pristine) and multi-walled (functionalized) CNTs are used as stabilizers to produce Pickering type oil-in-water (O/W) emulsions. Lipids namely, Dimodan U and Phytantriol are used as emulsifiers, which in excess water self-assemble into the bicontinuous cubic Pn3m phase. This highly viscous phase is fragmented into smaller particles using a probe ultrasonicator in presence of conventional surfactant stabilizers or CNTs as done here. Initially, the CNTs (powder form) are dispersed in water followed by further ultrasonication with the molten lipid to form the final emulsion. During this process the CNTs get coated with lipid molecules, which in turn are presumed to surround the lipid droplets to form a particulate emulsion that is stable for months. The average size of CNT-stabilized nanostructured lipid particles is in the submicron range, which compares well with the particles stabilized using conventional surfactants. Small angle X-ray scattering data confirms the retention of the original Pn3m cubic phase in the CNT-stabilized lipid dispersions as compared to the pure lipid phase (bulk state). Blue shift and lowering of the intensities in characteristic G and G&#39; bands of CNTs observed in Raman spectroscopy characterize the interaction between CNT surface and lipid molecules. These results suggest that the interactions between the CNTs and lipids are responsible for their mutual stabilization in aqueous solutions. As the concentrations of CNTs employed for stabilization are very low and lipid molecules are able to functionalize the CNTs, the toxicity of CNTs is expected to be insignificant while their biocompatibility is greatly enhanced. Hence the present approach finds a great potential in various biomedical applications, for instance, for developing hybrid nanocarrier systems for the delivery of multiple functional molecules as in combination therapy or polytherapy.

We report on a smart application of carbon nanotubes for kinetic stabilization of lipid particles that contain self-assembled nanostructures in their cores. The preparation of lipid particles requires rather low concentrations of carbon nanotubes permitting their use in biomedical applications such as drug delivery.

カーボンナノチューブにより安定化内部自己組織化脂質粒子の簡便な調製

We present a facile method to prepare nanostructured lipid particles stabilized by carbon nanotubes (CNTs). Single-walled (pristine) and multi-walled ...

Highly Stable, Functional Hairy Nanoparticles and Biopolymers from Wood Fibers: Towards Sustainable Nanotechnology

Nanoparticles, as one of the key materials in nanotechnology and nanomedicine, have gained significant importance during the past decade. While metal-based nanoparticles are associated with synthetic and environmental hassles, cellulose introduces a green, sustainable alternative for nanoparticle synthesis. Here, we present the chemical synthesis and separation procedures to produce new classes of hairy nanoparticles (bearing both amorphous and crystalline regions) and biopolymers based on wood fibers. Through periodate oxidation of soft wood pulp, the glucose ring of cellulose is opened at the C2-C3 bond to form 2,3-dialdehyde groups. Further heating of the partially oxidized fibers (e.g., T = 80 &#176;C) results in three products, namely fibrous oxidized cellulose, sterically stabilized nanocrystalline cellulose (SNCC), and dissolved dialdehyde modified cellulose (DAMC), which are well separated by intermittent centrifugation and co-solvent addition. The partially oxidized fibers (without heating) were used as a highly reactive intermediate to react with chlorite for converting almost all aldehyde to carboxyl groups. Co-solvent precipitation and centrifugation resulted in electrosterically stabilized nanocrystalline cellulose (ENCC) and dicarboxylated cellulose (DCC). The aldehyde content of SNCC and consequently surface charge of ENCC (carboxyl content) were precisely controlled by controlling the periodate oxidation reaction time, resulting in highly stable nanoparticles bearing more than 7 mmol functional groups per gram of nanoparticles (e.g., as compared to conventional NCC bearing &#60;&#60; 1 mmol functional group/g). Atomic force microscopy (AFM), transmission electron microscopy (TEM), and scanning electron microscopy (SEM) attested to the rod-like morphology. Conductometric titration, Fourier transform infrared spectroscopy (FTIR), nuclear magnetic resonance (NMR), dynamic light scattering (DLS), electrokinetic-sonic-amplitude (ESA) and acoustic attenuation spectroscopy shed light on the superior properties of these nanomaterials.

Synthesis schemes to prepare highly stable wood fiber-based hairy nanoparticles and functional cellulose-based biopolymers have been detailed.

高安定、機能毛深いナノ粒子と木材繊維からバイオポリマー：持続可能なナノテクノロジーに向けて

Nanoparticles, as one of the key materials in nanotechnology and nanomedicine, have gained significant importance during the past decade. While ...

Microfluidic-based Synthesis of Covalent Organic Frameworks (COFs): A Tool for Continuous Production of COF Fibers and Direct Printing on a Surface

Covalent Organic Frameworks (COFs) are a class of porous covalent materials which are frequently synthesized as unprocessable crystalline powders. The first COF was reported in 2005 with much effort centered on the establishment of new synthetic routes for its preparation. To date, most available synthetic methods for COF synthesis are based on bulk mixing under solvothermal conditions. Therefore, there is increasing interest in developing systematic protocols for COF synthesis that provide for fine control over reaction conditions and improve COF processability on surfaces, which is essential for their use in practical applications. Herein, we present a novel microfluidic-based method for COF synthesis where the reaction between two constituent building blocks, 1,3,5-benzenetricarbaldehyde (BTCA) and 1,3,5-tris(4-aminophenyl)benzene (TAPB), takes place under controlled diffusion conditions and at room temperature. Using such an approach yields sponge-like, crystalline fibers of a COF material, hereafter called MF-COF. The mechanical properties of MF-COF and the dynamic nature of the approach allow the continuous production of MF-COF fibers and their direct printing onto surfaces. The general method opens new potential applications requiring advanced printing of 2D or 3D COF structures on flexible or rigid surfaces.

Microfluidic-based Synthesis of Covalent Organic Frameworks COFs: A Tool for Continuous Production of COF Fibers and Direct Printing on a Surface

We present a novel microfluidic-based method for synthesis of covalent organic frameworks (COFs). We demonstrate how this approach can be used to produce continuous COF fibers, and also 2D or 3D COF structures on surfaces.

コバレント有機骨格（COF）のマイクロ流体ベースの合成：COF繊維の連続生産と表面への直接印刷のためのツール

Covalent Organic Frameworks (COFs) are a class of porous covalent materials which are frequently synthesized as unprocessable crystalline powders. The ...

Light-driven Molecular Motors on Surfaces for Single Molecular Imaging

The design and synthesis of a synthetic system that aims for the direct visualization of a synthetic rotary motor at the single molecule level on surfaces are demonstrated. This work requires careful design, considerable synthetic effort, and proper analysis. The rotary motion of the molecular motor in solution is shown by 1H NMR and UV-vis absorption spectroscopy techniques. In addition, the method to graft the motor onto an amine-coated quartz is described. This method helps to gain more insight into molecular machines.

The manuscript describes how to synthesize and graft a molecular motor on surfaces for single molecular imaging.

単一分子イメージングの表面上の光駆動の分子モーター

The design and synthesis of a synthetic system that aims for the direct visualization of a synthetic rotary motor at the single molecule level on surfaces ...

Analysis of Complex Molecules and Their Reactions on Surfaces by Means of Cluster-Induced Desorption/Ionization Mass Spectrometry

Desorption/Ionization Induced by Neutral SO2 Clusters (DINeC) is employed as a very soft and efficient desorption/ionization technique for mass spectrometry (MS) of complex molecules and their reactions on surfaces. DINeC is based on a beam of SO2 clusters impacting on the sample surface at low cluster energy. During cluster-surface impact, some of the surface molecules are desorbed and ionized via dissolvation in the impacting cluster; as a result of this dissolvation-mediated desorption mechanism, low cluster energy is sufficient and the desorption process is extremely soft. Both surface adsorbates and molecules of which the surface is composed of can be analyzed. Clear and fragmentation-free spectra from complex molecules such as peptides and proteins are obtained. DINeC does not require any special sample preparation, in particular no matrix has to be applied. The method yields quantitative information on the composition of the samples; molecules at a surface coverage as low as 0.1 % of a monolayer can be detected. Surface reactions such as H/D exchange or thermal decomposition can be observed in real-time and the kinetics of the reactions can be deduced. Using a pulsed nozzle for cluster beam generation, DINeC can be efficiently combined with ion trap mass spectrometry. The matrix-free and soft nature of the DINeC process in combination with the MSn capabilities of the ion trap allows for very detailed and unambiguous analysis of the chemical composition of complex organic samples and organic adsorbates on surfaces.

Neutral SO2 clusters of low kinetic energy (&#60; 0.8 eV/constituent) are used to desorb complex surface molecules such as peptides or lipids for further analysis by means of mass spectrometry using an ion trap mass spectrometer. No special sample preparation is required, and real-time observation of reactions is possible.

クラスターによる脱着/イオン化質量分析による、表面上の複雑分子とその反応の解析

Desorption/Ionization Induced by Neutral SO2 Clusters (DINeC) is employed as a very soft and efficient desorption/ionization technique for mass ...

Covalent Attachment of Single Molecules for AFM-based Force Spectroscopy

Atomic force microscopy (AFM)-based single molecule force spectroscopy is an ideal tool for investigating the interactions between a single polymer and surfaces. For a true single molecule experiment, covalent attachment of the probe molecule is essential because only then can hundreds of force-extension traces with one and the same single molecule be obtained. Many traces are in turn necessary to prove that a single molecule alone is probed. Additionally, passivation is crucial for preventing unwanted interactions between the single probe molecule and the AFM cantilever tip as well as between the AFM cantilever tip and the underlying surface. The functionalization protocol presented here is reliable and can easily be applied to a variety of polymers. Characteristic single molecule events (i.e., stretches and plateaus) are detected in the force-extension traces. From these events, physical parameters such as stretching force, desorption force and desorption length can be obtained. This is particularly important for the precise investigation of stimuli-responsive systems at the single molecule level. As exemplary systems poly(ethylene glycol) (PEG), poly(N-isopropylacrylamide) (PNiPAM) and polystyrene (PS) are stretched and desorbed from SiOx (for PEG and PNiPAM) and from hydrophobic self-assembled monolayer surfaces (for PS) in aqueous environment.

Covalent attachment of probe molecules to atomic force microscopy (AFM) cantilever tips is an essential technique for the investigation of their physical properties. This allows us to determine the stretching force, desorption force and length of polymers via AFM-based single molecule force spectroscopy with high reproducibility.

AFMベースの力分光法のための単一分子の共有結合

Atomic force microscopy (AFM)-based single molecule force spectroscopy is an ideal tool for investigating the interactions between a single polymer and ...

Synthesis of pH Dependent Pyrazole, Imidazole, and Isoindolone Dipyrrinone Fluorophores using a Claisen-Schmidt Condensation Approach

Methine-bridged conjugated bicyclic aromatic compounds are common constituents of a range of biologically relevant molecules such as porphyrins, dipyrrinones, and pharmaceuticals. Additionally, restricted rotation of these systems often results in highly to moderately fluorescent systems as observed in 3H,5H-dipyrrolo[1,2-c:2',1'-f]pyrimidin-3-ones, xanthoglows, pyrroloindolizinedione analogs, BODIPY analogs, and the phenolic and imidazolinone ring systems of Green Fluorescent Protein (GFP). This manuscript describes an inexpensive and operationally simple method of performing a Claisen-Schmidt condensation to generate a series of fluorescent pH dependent pyrazole/imidazole/isoindolone dipyrrinone analogs. While the methodology illustrates the synthesis of dipyrrinone analogs, it can be translated to produce a wide range of conjugated bicyclic aromatic compounds. The Claisen-Schmidt condensation reaction utilized in this method is limited in scope to nucleophiles and electrophiles that are enolizable under basic conditions (nucleophile component) and non-enolizable aldehydes (electrophile component). Additionally, both the nucleophilic and electrophilic reactants must contain functional groups that will not inadvertently react with hydroxide. Despite these limitations, this methodology offers access to completely novel systems that can be employed as biological or molecular probes.

The Claisen-Schmidt condensation reaction is an important methodology for the generation of methine-bridged conjugated bicyclic aromatic compounds. Through utilizing a base-mediated variant of the aldol reaction, a range of fluorescent and/or biologically relevant molecules can be accessed through a generally inexpensive and operationally simple synthetic approach.

クレイゼン・シュミット凝縮アプローチによるpH依存性ピラゾール、イミダゾール、イソインドロネジピロンフルオロフォアの合成

Methine-bridged conjugated bicyclic aromatic compounds are common constituents of a range of biologically relevant molecules such as porphyrins, ...

Analysis of Organochlorine Pesticides in a Soil Sample by a Modified QuEChERS Approach Using Ammonium Formate

Currently, the QuEChERS method represents the most widely used sample preparation protocol worldwide for analyzing pesticide residues in a broad variety of matrices both in official and non-official laboratories. The QuEChERS method using ammonium formate has previously proven to be advantageous compared to the original and the two official versions. On the one hand, the simple addition of 0.5 g of ammonium formate per gram of sample is sufficient to induce phase separation and achieve good analytical performance. On the other hand, ammonium formate reduces the need for maintenance in routine analyses. Here, a modified QuEChERS method using ammonium formate was applied for the simultaneous analysis of organochlorine pesticide (OCP) residues in agricultural soil. Specifically, 10 g of the sample was hydrated with 10 mL of water and then extracted with 10 mL of acetonitrile. Next, phase separation was carried out using 5 g of ammonium formate. After centrifugation, the supernatant was subjected to a dispersive solid-phase extraction clean-up step with anhydrous magnesium sulfate, primary-secondary amine, and octadecylsilane. Gas chromatography-mass spectrometry was used as the analytical technique. The QuEChERS method using ammonium formate is demonstrated as a successful alternative for extracting OCP residues from a soil sample.

The present protocol describes the utilization of ammonium formate for phase partitioning in QuEChERS, together with gas chromatography-mass spectrometry, to successfully determine organochlorine pesticide residues in a soil sample.

ギ酸アンモニウムを用いた改変QuEChERS法による土壌試料中の有機塩素系農薬の分析

Currently, the QuEChERS method represents the most widely used sample preparation protocol worldwide for analyzing pesticide residues in a broad variety ...