1。脂質の準備<ol><li>二重層の上部のリーフレットにタンパク質をリンクするために、我々は、Ni 2を使用してください+キレート脂質、ポリヒスチジンタグを持つアンカー組換えタンパク質を。特定の細胞型は、Ni 2 +でコーティングされた二重層へ非特異的に付着する可能性がありますが、T細胞は、一般的にT細胞活性化を研究するためのこの方法が最適なこと、しないでください。 </li><li>脂質二重層液をクロロホルム-メタノールの1〜2 mlのガラス管に適当な量のNi 2 +キレート脂質と80％のホスファチジルコリン20％の溶液を調製することによって行われます。 <ol><li>暖かい（30〜37℃）の水浴中で窒素ガス気流下で溶媒を蒸発させる。これは通常、約15分かかります。 </li><li> 90分間凍結乾燥機を使用して、高真空下で、残りの溶剤を削除します。 </li></ol></li><li>混合界面活性剤/リン脂質ミセルのソリューションを作成するトリス - 生理食塩水緩衝液、2％N -オクチル-グルコシドの洗剤で脂質を溶解する。最終的なリン脂質濃度は、0.4mMのはずです。リポソームを形成するために、洗剤とフォームリポソームを削除するには、トリス - 生理食塩水を3変更を避けるために、このソリューションを透析。我々は通常、10 kDaの周りのカットオフを持つスペクトラ/ POR第2配管直径6mm 1 mlの順に容量ボリュームで機能します。我々は、チューブをクランプするとき、任意の気泡のを除外するように注意です。我々は典型的には、滅菌フィルターこのソリューションおよび70％エタノール、滅菌手袋を使用してそれを処理し、クリーンな条件下での透析を行う。 </li><li>このソリューションでは、水晶明確にする必要があります。それは酸化を防ぐために、アルゴンガス下に格納されています。解決策が濁っている場合は、多層小胞が生成されている、と再度起動する必要があります。 </li></ol> 2。 ICAM - 1とMHCは平面二重膜上に堆積されているどのくらいの決定<ol><li>実験をセットアップするためには、可溶性タンパク質の特定の濃度でのNi 2 +キレート二重層の特定の表面積上に堆積されているどのくらいのICAM - 1とMHCを決定するためにまず必要です。 5マイクロメートル径のガラスビーズでの預金二重層、これを行うには、イメージングに使用されるフ​​ローの細胞のもの、およびフロー免疫蛍光アッセイによる結合タンパク質の密度を読み出すおおよそその条件下でのタンパク質溶液とガラスビーズをインキュベートする。 </li><li>フルオレセイン分子当たりの知られている番号とFITC標識抗体を表面に結合したタンパク質を検出するために使用されます。フルオレセイン標準ビーズをフローmicrofluorimeterをキャリブレーションに使用されます。 </li><li>我々は、I - EK - MCC91 - 103複合体のICAM - 1の200molecules/μm2と0.2から20molecules/μm2と抗原提示細胞上のものに近似する条件を確立します。 </li></ol> 3。平面脂質二重層を形成するためのガラスのカバースリップのクリーニング<ol><li>平面脂質二重層は、リポソームがガラスに融合するときに自発的に形成するが、ガラスが適切に洗浄されている場合のみ。 </li><li>ピラニアで作業する場合、我々は酸のエプロンと重い、耐酸性の籠手を含む、完全な防護服を着用してください。慎重に、有機廃棄物から廃棄物を分離し、適切に処分する。 </li><li>酸のピラニア溶液を調製するために、乾燥したビーカーに濃硫酸75mlのを追加し、慎重に30％過酸化水素25mlを加える。ソリューションは、急速に超えて100℃に加熱</li><li> 15分間溶液中で乾燥したカバーグラスを浸し。ソリューションから削除し、それぞれの側分間精製水ですすいでください。精製水を切るために真空源を用いてカバースリップを乾かします。ピラニア溶液を冷却し、よくドラフト内でマークキャップ緩み、が残っているピラニアの廃棄物コンテナに追加することができます。リポソーム、カバーガラス、及びタンパク質のすべてが準備されている場合、免疫シナプスが形成される準備ができています。 </li></ol> 4。二重層を形成する<ol><li>脂質二重層を形成するために、私たちの研究室では、統合された加熱の利点を持っているBioptechs FCSIIチャンバーを、使用しています。 FCSIIシステムは、ステンレス製のベースになりますが一緒にクランプの反対側の流体溝と0.5ミリメートルトップガスケット、一方の表面に加熱するためのインジウムスズ酸化物で被覆microaqueductスライド、マイクロ流体マニホールド、ダストフリー0.25ミリメートルフローチャンバーとピラニアの高さを定義するガスケットは、二重層が形成されている40mmの円形のカバースリップを掃除した。 </li><li>二重層が形成される際に、カバースリップは、親水性の高い、クリーンにする必要がある一方でそれはより疎水性なら、microacqueductスライドは、実際にうまく機能します。 microaqueductを作ることはより多くの疎水性は、純水で洗浄し、この調整のプロセスの後、同様の用途の間で完全に乾燥、室温で60分間、1％HSAで処理することにより達成されます。この手順によって残された変性タンパク質のコーティングは、リポソーム懸濁液を1μl、ラウンド、hemisphericaを形成することができます&gt; 0.25 mmの高であるlのドロップ。 </li><li>フローセルとフォーム平面二重膜を組み立てるには、平らな表面上に、上向きに指示チューブで、マニホールドを配置。その後、0を置きます。それが平らに装着さ​​れていることを確認して、流体管の上に置く穴と5 mmのガスケット。ゆっくりと上向きに流体チャンネルとガスケットにmicroaqueduct側に配置し、スライドの穴がマイクロ流体管に合わせていることを検証するまでは、どんな下向きの圧力を適用せずにフラットシートを確認してください。チャネルが流体チャネルと揃うとは大きな空気のスペースがないことになるようにmicroaqueductスライドの上から長方形のカットでダストフリー0.25mmのガスケットを置きます。 </li><li>各ドロップの中心間距離が2.5mmになるようなパターンでmicroaqueductスライドの表面に5にリポソーム懸濁液のX1μlの水滴を置きます。 5リポソーム滴は異なる組成を持つことができますが、この場合、我々は2つだけ二層、ニッケル2 + +キレート脂質および10％のNi 2 + +のキレート脂質とつのNOを持つものを形成します。 </li><li>これらのドロップが正しく設定されると、慎重にカバーが一つの動きの面を下にスリップ置きます。できるだけ早くベースの上にクランプ。リポソーム滴はカバースリップとの接触を確認する必要があります。脂質二重層は、おそらくすでに形成されているので、この接点は、異常はありません。すべての中断は、欠陥のある二層になることがあります。 </li><li>顕微鏡のカバーの位置決めを支援するためにmicroacqueductスライド上のいくつかの小さなインクスポットのある二層の位置をマークするために覚えておくことが重要です。システムが平衡しているか確かめる為に、20分待ってください。約20フレキシブルチューブのcmおよび3方活栓に接続した20mlの注射器を使用して、すべてのソリューションを転送します。コックとフローセルの間にデッドボリュームを最小限に抑えるためにチューブの短い長さでフローセルに活栓の別のポートを接続します。 2mMのMgCl 2、1mMのCaCl 2、5mMグルコースと1％ヒト血清アルブミンを含む生理食塩水HEPES緩衝でシリンジを埋める。 </li><li>に気泡がないので素数ではチューブとコック。フローセルにこれを接続していない空気の泡が二重層を通過しないことを確認するために、見ながら一つの動きで5ミリリットルを介してプッシュ。今、あなたの二重膜を持つことになります。 </li></ol>条件：このビデオの記事は&quot;アメーバロコモーションの主題による変奏曲など、免疫学的シナプスとKinapsesギャザラと戻るハンター&quot;サポートされています<a href="http://arjournals.annualreviews.org/action/doSearch?action=runSearch&amp;type=advanced&amp;result=true&amp;prevSearch=%2Bauthorsfield%3A%28Dustin%2C+Michael+L.%29" target="_blank">マイケルL.ダスティン</a> 、 <a href="http://arjournals.annualreviews.org/doi/abs/10.1146/annurev.cellbio.24.110707.175226">細胞および発生生物学の年次レビュー</a> 、ボリューム24、2008。

supported planar bilayers for the formation of study of immunological synapses and kinapse

サポートされている平面脂質二重層は、免疫学的シナプスの分子間相互作用をモデル化するために使用できる強力なツールです。リンパ球と抗原提示細胞間の相互作用を模倣するために、我々は、ポリ - ヒスチジンタグを持つ二層の上部のリーフレットに組換え細胞接着およびMHCタンパク質を固定するNI2 +キレート脂質を使用してください。平面脂質二重層は、脂質の準備二重層が形成されるガラスの表面を処理し、リンパ球が追加されるフローセルを含む特殊なチャンバー内の二重層を形成することによって生成されます。その後、二層をNiで起訴され、Hisタグ融合組み換えタンパク質が追加されます。最後に、リンパ球は、フローセルに注入され、全反射顕微鏡は、画像のシナプス形成とメカニズム、その制御性T細胞の移動運動、受容体のソートのサイト、および受容体の劣化の部位に使用することができます。

Watch this Scientific Journal Video about Supported Planar Bilayers for the Formation of Study of Immunological Synapses and Kinapse at JoVE.com

Supported Planar Bilayers for the Formation of Study of Immunological Synapses and Kinapse

サポートされている平面脂質二重層は、免疫学的シナプスの分子間相互作用をモデル化するために使用できる強力なツールです。ここで、我々は、脂質のbilyerの上部のリーフレットにシナプス形成を調節し、全反射顕微鏡を用いてシナプス形成を可視化するために知られている細胞接着タンパク質を固定するための方法を示しています。

免疫シナプスとKinapseの研究の形成のためのサポートされている平面二重層

Supported planar bilayers are powerful tools that can be used to model the molecular interactions in an immunological synapse.   To mimic the interactions ...

supported-planar-bilayers-for-formation-study-immunological-synapses

Research

JoVE Journal

Biology

12.5K ビュー.  New York University - NYU. サポートされている平面脂質二重層は、免疫学的シナプスの分子間相互作用をモデル化するために使用できる強力なツールです。ここで、我々は、脂質のbilyerの上部のリーフレットにシナプス形成を調節し、全反射顕微鏡を用いてシナプス形成を可視化するために知られている細胞接着タンパク質を固定するための方法を示しています。

ビデオ: Supported Planar Bilayers for the Formation of Study of Immunological Synapses and Kinapse

Testing Visual Sensitivity to the Speed and Direction of Motion in Lizards

Testing visual sensitivity in any species provides basic information regarding behaviour, evolution, and ecology. However, testing specific features of the visual system provide more empirical evidence for functional applications. Investigation into the sensory system provides information about the sensory capacity, learning and memory ability, and establishes known baseline behaviour in which to gauge deviations (Burghardt, 1977). However, unlike mammalian or avian systems, testing for learning and memory in a reptile species is difficult. Furthermore, using an operant paradigm as a psychophysical measure of sensory ability is likewise as difficult. Historically, reptilian species have responded poorly to conditioning trials because of issues related to motivation, physiology, metabolism, and basic biological characteristics. Here, I demonstrate an operant paradigm used a novel model lizard species, the Jacky dragon (Amphibolurus muricatus) and describe how to test peripheral sensitivity to salient speed and motion characteristics. This method uses an innovative approach to assessing learning and sensory capacity in lizards. I employ the use of random-dot kinematograms (RDKs) to measure sensitivity to speed, and manipulate the level of signal strength by changing the proportion of dots moving in a coherent direction. RDKs do not represent a biologically meaningful stimulus, engages the visual system, and is a classic psychophysical tool used to measure sensitivity in humans and other animals. Here, RDKs are displayed to lizards using three video playback systems. Lizards are to select the direction (left or right) in which they perceive dots to be moving. Selection of the appropriate direction is reinforced by biologically important prey stimuli, simulated by computer-animated invertebrates.

Testing visual sensitivity in lizards using an operant conditioning paradigm that employs video playback of random-dot kinematograms and computer-generated invertebrates.

トカゲの運動の速度と方向の視覚感度のテスト

Testing visual sensitivity in any species provides basic information regarding behaviour, evolution, and ecology. However, testing specific features of ...

生物学

Computer-Generated Animal Model Stimuli

Communication between animals is diverse and complex. Animals may communicate using auditory, seismic, chemosensory, electrical, or visual signals. In particular, understanding the constraints on visual signal design for communication has been of great interest. Traditional methods for investigating animal interactions have used basic observational techniques, staged encounters, or physical manipulation of morphology. Less intrusive methods have tried to simulate conspecifics using crude playback tools, such as mirrors, still images, or models. As technology has become more advanced, video playback has emerged as another tool in which to examine visual communication (Rosenthal, 2000). However, to move one step further, the application of computer-animation now allows researchers to specifically isolate critical components necessary to elicit social responses from conspecifics, and manipulate these features to control interactions. Here, I provide detail on how to create an animation using the Jacky dragon as a model, but this process may be adaptable for other species. In building the animation, I elected to use Lightwave  3D to alter object morphology, add texture, install bones, and provide comparable weight shading that prevents exaggerated movement. The animation is then matched to select motor patterns to replicate critical movement features. Finally, the sequence must rendered into an individual clip for presentation. Although there are other adaptable techniques, this particular method had been demonstrated to be effective in eliciting both conspicuous and social responses in staged interactions.

Computer-generated stimuli using the Jacky dragon as a model.

コンピュータによって生成された動物モデルの刺激

Communication between animals is diverse and complex. Animals may communicate using auditory, seismic, chemosensory, electrical, or visual signals. In ...

Investigating the Immunological Mechanisms Underlying Organ Transplant Rejection

免疫学的メカニズム深部臓器移植の拒絶反応の調査

Preparation of Artificial Bilayers for Electrophysiology Experiments

Planar lipid bilayers, also called artificial lipid bilayers, allow you to study ion-conducting channels in a well-defined environment. These bilayers can be used for many different studies, such as the characterization of membrane-active peptides, the reconstitution of ion channels or investigations on how changes in lipid bilayer properties alter the function of bilayer-spanning channels.  Here, we show how to form a planar bilayer and how to isolate small patches from the bilayer, and in a second video  will also demonstrate a procedure for using gramicidin channels to determine changes in lipid bilayer elastic properties.   We also demonstrate the individual steps needed to prepare the bilayer chamber, the electrodes and how to test that the bilayer is suitable for single-channel measurements.

Planar lipid bilayers, also called artificial lipid bilayers, allow you to study ion-conducting channels in a well-defined environment. Here, we demonstrate the individual steps needed to prepare the bilayer chamber, the electrodes and how to test that the bilayer is suitable for single-channel measurements.

電気生理学的実験のための人工二重層の準備

Planar lipid bilayers, also called artificial lipid bilayers, allow you to study ion-conducting channels in a well-defined environment. These bilayers can ...

Isolation and Purification of Drosophila Peripheral Neurons by Magnetic Bead Sorting

The Drosophila peripheral nervous system (PNS) is a powerful model for investigating the complex processes of neuronal development and dendrite morphogenesis at the functional and molecular levels. To aid in these analyses, we have developed a strategy for the isolation of a subclass of PNS neurons called dendritic arborization (da) neurons that have been widely used for studying dendrite morphogenesis1,2. These neurons are very difficult to isolate as a pure population, due in part to their extremely low occurrence and their difficult-to-reach location below the tough chitinous larval cuticle. Our newly developed method overcomes these challenges, and is based on a fast and specific cell enrichment using antibody-coated magnetic beads. For our magnetic bead sorting studies, we have used age-matched third instar larvae expressing a mouse CD8 tagged GFP fusion protein (UAS-mCD8-GFP)3 under the control of either the class IV dendritic arborization (da) neuron-specific pickpocket (ppk)-GAL4 driver4 or the control of the pan-da neuron-specific GAL421-7 driver5. Although this protocol has been optimized for isolating PNS cells which are attached to the inner wall of the larval cuticle, by varying a few parameters, the same protocol could be used to isolate many different cell types attached to the cuticle at larval or pupal stages of development (e.g. epithelia, muscle, oenocytes etc.), or other cell types from larval organs depending upon the GAL4-specific driver expression pattern. The RNA isolated by this method is of high quality and can be readily used for downstream genomic analyses such as microarray gene expression profiling studies. This approach offers a powerful new tool to perform studies on isolated Drosophila dendritic arborization (da) neurons thereby providing novel insights into the molecular mechanisms underlying dendrite morphogenesis.

Isolation and Purification of Drosophila Peripheral Neurons by Magnetic Bead Sorting

In this video-article we present a method for the isolation and purification of Drosophila peripheral neurons using a fast magnetic bead assisted cell sorting strategy. RNA obtained from the isolated cells can be readily used for downstream applications including microarray analyses.

の単離と精製ショウジョウバエ</em磁気ビーズソート別>末梢ニューロン

The Drosophila peripheral nervous system (PNS) is a powerful model for investigating the complex processes of neuronal development and dendrite ...

Do-It-Yourself Device for Recovery of Cryopreserved Samples Accidentally Dropped into Cryogenic Storage Tanks

Liquid nitrogen is colorless, odorless, extremely cold (-196 &deg;C) liquid kept under pressure. It is commonly used as a cryogenic fluid for long term storage of biological materials such as blood, cells and tissues 1,2. The cryogenic nature of liquid nitrogen, while ideal for sample preservation, can cause rapid freezing of live tissues on contact - known as 'cryogenic burn'2, which may lead to severe frostbite in persons closely involved in storage and retrieval of samples from Dewars. Additionally, as liquid nitrogen evaporates it reduces the oxygen concentration in the air and might cause asphyxia, especially in confined spaces2.
In laboratories, biological samples are often stored in cryovials or cryoboxes stacked in stainless steel racks within the Dewar tanks1. These storage racks are provided with a long shaft to prevent boxes from slipping out from the racks and into the bottom of Dewars during routine handling. All too often, however, boxes or vials with precious samples slip out and sink to the bottom of liquid nitrogen filled tank. In such cases, samples could be tediously retrieved after transferring the liquid nitrogen into a spare container or discarding it. The boxes and vials can then be relatively safely recovered from emptied Dewar. However, the cryogenic nature of liquid nitrogen and its expansion rate makes sunken sample retrieval hazardous. It is commonly recommended by Safety Offices that sample retrieval be never carried out by a single person. Another alternative is to use commercially available cool grabbers or tongs to pull out the vials3. However, limited visibility within the dark liquid filled Dewars poses a major limitation in their use.
In this article, we describe the construction of a Cryotolerant DIY retrieval device, which makes sample retrieval from Dewar containing cryogenic fluids both safe and easy.

Here we present a low cost, durable cryotolerant device for sample retrieval from Dewar tanks filled with liquid nitrogen. The ease of construction and modular design of the device makes the process of sample retrieval from cryogenic tanks safe and easy.

凍結保存サンプルの回復のための日曜大工のデバイスが誤って低温貯蔵タンクに落とし

Liquid nitrogen is colorless, odorless, extremely cold (-196 &deg;C) liquid kept under pressure. It is commonly used as a cryogenic fluid for long term ...

Introduction to Solid Supported Membrane Based Electrophysiology

The electrophysiological method we present is based on a solid supported membrane (SSM) composed of an octadecanethiol layer chemisorbed on a gold coated sensor chip and a phosphatidylcholine monolayer on top. This assembly is mounted into a cuvette system containing the reference electrode, a chlorinated silver wire.
After adsorption of membrane fragments or proteoliposomes containing the membrane protein of interest, a fast solution exchange is used to induce the transport activity of the membrane protein. In the single solution exchange protocol two solutions, one non-activating and one activating solution, are needed. The flow is controlled by pressurized air and a valve and tubing system within a faraday cage.
The kinetics of the electrogenic transport activity is obtained via capacitive coupling between the SSM and the proteoliposomes or membrane fragments. The method, therefore, yields only transient currents. The peak current represents the stationary transport activity. The time dependent transporter currents can be reconstructed by circuit analysis.
This method is especially suited for prokaryotic transporters or eukaryotic transporters from intracellular membranes, which cannot be investigated by patch clamp or voltage clamp methods.

Here we present an electrophysiological method based on solid supported membranes with focus on its applications for the characterization of electrogenic membrane transporters.

電気生理学基づき固体支持された膜の概要

The electrophysiological method we present is based on a solid supported membrane (SSM) composed of an octadecanethiol layer chemisorbed on a gold coated ...

Direct Imaging of ER Calcium with Targeted-Esterase Induced Dye Loading (TED)

Visualization of calcium dynamics is important to understand the role of calcium in cell physiology. To examine calcium dynamics, synthetic fluorescent Ca2+ indictors have become popular. Here we demonstrate TED (= targeted-esterase induced dye loading), a method to improve the release of Ca2+ indicator dyes in the ER lumen of different cell types. To date, TED was used in cell lines, glial cells, and neurons in vitro. TED bases on efficient, recombinant targeting of a high carboxylesterase activity to the ER lumen using vector-constructs that express Carboxylesterases (CES). The latest TED vectors contain a core element of CES2 fused to a red fluorescent protein, thus enabling simultaneous two-color imaging. The dynamics of free calcium in the ER are imaged in one color, while the corresponding ER structure appears in red. At the beginning of the procedure, cells are transduced with a lentivirus. Subsequently, the infected cells are seeded on coverslips to finally enable live cell imaging. Then, living cells are incubated with the acetoxymethyl ester (AM-ester) form of low-affinity Ca2+ indicators, for instance Fluo5N-AM, Mag-Fluo4-AM, or Mag-Fura2-AM. The esterase activity in the ER cleaves off hydrophobic side chains from the AM form of the Ca2+ indicator and a hydrophilic fluorescent dye/Ca2+ complex is formed and trapped in the ER lumen. After dye loading, the cells are analyzed at an inverted confocal laser scanning microscope. Cells are continuously perfused with Ringer-like solutions and the ER calcium dynamics are directly visualized by time-lapse imaging. Calcium release from the ER is identified by a decrease in fluorescence intensity in regions of interest, whereas the refilling of the ER calcium store produces an increase in fluorescence intensity. Finally, the change in fluorescent intensity over time is determined by calculation of &Delta;F/F0.

Direct Imaging of ER Calcium with Targeted-Esterase Induced Dye Loading TED

Targeted-esterase induced dye loading (TED) supports the analysis of intracellular calcium store dynamics by fluorescence imaging. The method bases on targeting of a recombinant Carboxylesterase to the endoplasmic reticulum (ER), where it improves the local unmasking of synthetic low-affinity Ca2+ indicator dyes in the ER lumen.

ターゲット-エステラーゼ誘起染料の読み込み（TED）とERカルシウムの直接イメージング

Visualization of calcium dynamics is important to understand the role of calcium in cell physiology. To examine calcium dynamics, synthetic fluorescent ...

Endothelial Cell Tube Formation Assay for the In Vitro Study of Angiogenesis

Angiogenesis is a vital process for normal tissue development and wound healing, but is also associated with a variety of pathological conditions. Using this protocol, angiogenesis may be measured in vitro in a fast, quantifiable manner. Primary or immortalized endothelial cells are mixed with conditioned media and plated on basement membrane matrix. The endothelial cells form capillary like structures in response to angiogenic signals found in conditioned media. The tube formation occurs quickly with endothelial cells beginning to align themselves within 1&#160;hr and lumen-containing tubules beginning to appear within 2 hr. Tubes can be visualized using a phase contrast inverted microscope, or the cells can be treated with calcein AM prior to the assay and tubes visualized through fluorescence or confocal microscopy. The number of branch sites/nodes, loops/meshes, or number or length of tubes formed can be easily quantified as a measure of in vitro angiogenesis. In summary, this assay can be used to identify genes and pathways that are involved in the promotion or inhibition of angiogenesis in a rapid, reproducible, and quantitative manner.

Endothelial Cell Tube Formation Assay for the In Vitro Study of Angiogenesis

The tube formation assay is a fast, quantifiable method for measuring in vitro angiogenesis. Endothelial cells are combined with conditioned media and plated on basement membrane extract. Tube formation occurs within hours and newly formed tubules easily quantified.

のための内皮細胞チューブ形成アッセイ<em&gt;体外</em&gt;血管新生に関する研究

Angiogenesis is a vital process for normal tissue development and wound healing, but is also associated with a variety of pathological conditions. Using ...

High-throughput Image Analysis of Tumor Spheroids: A User-friendly Software Application to Measure the Size of Spheroids Automatically and Accurately

The increasing number of applications of three-dimensional (3D) tumor spheroids as an in vitro model for drug discovery requires their adaptation to large-scale screening formats in every step of a drug screen, including large-scale image analysis. Currently there is no ready-to-use and free image analysis software to meet this large-scale format. Most existing methods involve manually drawing the length and width of the imaged 3D spheroids, which is a tedious and time-consuming process. This study presents a high-throughput image analysis software application &#8211; SpheroidSizer, which measures the major and minor axial length of the imaged 3D tumor spheroids automatically and accurately; calculates the volume of each individual 3D tumor spheroid; then outputs the results in two different forms in spreadsheets for easy manipulations in the subsequent data analysis. The main advantage of this software is its powerful image analysis application that is adapted for large numbers of images. It provides high-throughput computation and quality-control workflow. The estimated time to process 1,000 images is about 15 min&#160;on a minimally configured laptop, or around 1 min&#160;on a multi-core performance workstation. The graphical user interface (GUI) is also designed for easy quality control, and users can manually override the computer results. The key method used in this software is adapted from the active contour algorithm, also known as Snakes, which is especially suitable for images with uneven illumination and noisy background that often plagues automated imaging processing in high-throughput screens. The complimentary &#8220;Manual Initialize&#8221; and &#8220;Hand Draw&#8221; tools provide the flexibility to SpheroidSizer in dealing with various types of spheroids and diverse quality images. This high-throughput image analysis software remarkably reduces labor and speeds up the analysis process. Implementing this software is beneficial for 3D tumor spheroids to become a routine in vitro model for drug screens in industry and academia.

We present a high-throughput image analysis software application to measure the size of three-dimensional tumor spheroids imaged with bright-field microscopy. This application provides a fast and effective way to examine the effects of therapeutic drugs on spheroids, which is beneficial for researchers who wish to use spheroids in drug screens.

腫瘍球の高スループット画像解析：自動的かつ正確にスフェロイドのサイズを測定するための使いやすいソフトウェアアプリケーション

The increasing number of applications of three-dimensional (3D) tumor spheroids as an in vitro model for drug discovery requires their adaptation to ...