著者はリサの都市の優秀な技術的な支援をありがちましょう。

<ol>
	<li>Aigner, B., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Transgenic+pigs+as+models+for+translational+biomedical+research.">Transgenic pigs as models for translational biomedical research.</a> J Mol. Med. 88, 653-664 (2010).</li><li>Blutke, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Munich+MIDY+Pig+Biobank:+A+unique+resource+for+studying+organ+crosstalk+in+diabetes.">The Munich MIDY Pig Biobank: A unique resource for studying organ crosstalk in diabetes.</a> Mol Metab. 6, 931-940 (2017).</li><li>Klymiuk, N., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dystrophin-deficient+pigs+provide+new+insights+into+the+hierarchy+of+physiological+derangements+of+dystrophic+muscle.">Dystrophin-deficient pigs provide new insights into the hierarchy of physiological derangements of dystrophic muscle.</a> Hum Mol Genet. 22, 4368-4382 (2013).</li><li>Klymiuk, N., Seeliger, F., Bohlooly, Y. M., Blutke, A., Rudmann, D. G., Wolf, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Tailored+pig+models+for+preclinical+efficacy+and+safety+testing+of+targeted+therapies.">Tailored pig models for preclinical efficacy and safety testing of targeted therapies.</a> Toxicol Pathol. 44, 346-357 (2016).</li><li>Renner, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Permanent+neonatal+diabetes+in+INSC94Y+transgenic+pigs.">Permanent neonatal diabetes in INSC94Y transgenic pigs.</a> Diabetes. 62, 1505-1511 (2013).</li><li>Abbott, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Inside+the+first+pig+biobank.">Inside the first pig biobank.</a> Nature. 519, 397-398 (2015).</li><li>Albl, B., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Tissue+sampling+guides+for+porcine+biomedical+models.">Tissue sampling guides for porcine biomedical models.</a> Toxicol Pathol. 44, 414-420 (2016).</li><li>Gundersen, H. J. G., Mirabile, R., Brown, D., Boyce, R. W., Haschek, W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Stereological+principles+and+sampling+procedures+for+toxicologic+pathologists.">Stereological principles and sampling procedures for toxicologic pathologists.</a> Haschek and Rousseaux's Handbook of Toxicologic Pathology. , 215-286 (2013).</li><li>Howard, C. V., Reed, M. 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G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+isector:+A+simple+and+direct+method+for+generating+isotropic,+uniform+random+sections+from+small+specimens.">The isector: A simple and direct method for generating isotropic, uniform random sections from small specimens.</a> J Microsc. 165, 427-431 (1992).</li><li>Tschanz, S., Schneider, J. P., Knudsen, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Design-based+stereology:+Planning,+volumetry+and+sampling+are+crucial+steps+for+a+successful+study.">Design-based stereology: Planning, volumetry and sampling are crucial steps for a successful study.</a> Ann Anat. 196, 3-11 (2014).</li><li>Dorph-Petersen, K. A., Nyengaard, J. R., Gundersen, H. 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B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+efficiency+of+systematic+sampling+in+stereology+and+its+prediction.">The efficiency of systematic sampling in stereology and its prediction.</a> J Microsc. 147, 229-263 (1987).</li><li>Scherle, W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+simple+method+for+volumetry+of+organs+in+quantitative+stereology.">A simple method for volumetry of organs in quantitative stereology.</a> Mikroskopie. 26, 57-60 (1970).</li><li>Nielsen, K. K., Andersen, C. B., Kromann-Andersen, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+comparison+between+the+effects+of+paraffin+and+plastic+embedding+of+the+normal+and+obstructed+minipig+detrusor+muscle+using+the+optical+disector.">A comparison between the effects of paraffin and plastic embedding of the normal and obstructed minipig detrusor muscle using the optical disector.</a> J Urol. 154, 2170-2173 (1995).</li><li>Schneider, J. P., Ochs, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Alterations+of+mouse+lung+tissue+dimensions+during+processing+for+morphometry:+a+comparison+of+methods.">Alterations of mouse lung tissue dimensions during processing for morphometry: a comparison of methods.</a> Am J Physiol Lung Cell Mol Physiol. 306, L341-L350 (2014).</li><li>von Bartheld, C. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Distribution+of+particles+in+the+z-axis+of+tissue+sections:+Relevance+for+counting+methods.">Distribution of particles in the z-axis of tissue sections: Relevance for counting methods.</a> NeuroQuantology. 10, 66-75 (2012).</li><li>Baddeley, A. J., Gundersen, H. J., Cruz-Orive, L. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Estimation+of+surface+area+from+vertical+sections.">Estimation of surface area from vertical sections.</a> J microsc. 142, 259-276 (1986).</li><li>Blutke, A., Schneider, M. R., Wolf, E., Wanke, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Growth+hormone+(GH)-transgenic+insulin-like+growth+factor+1+(IGF1)-deficient+mice+allow+dissociation+of+excess+GH+and+IGF1+effects+on+glomerular+and+tubular+growth.">Growth hormone (GH)-transgenic insulin-like growth factor 1 (IGF1)-deficient mice allow dissociation of excess GH and IGF1 effects on glomerular and tubular growth.</a> Physiol Rep. 4, e12709 (2016).</li><li>Hermanns, W., Liebig, K., Schulz, L. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Postembedding+immunohistochemical+demonstration+of+antigen+in+experimental+polyarthritis+using+plastic+embedded+whole+joints.">Postembedding immunohistochemical demonstration of antigen in experimental polyarthritis using plastic embedded whole joints.</a> Histochemistry. 73, 439-446 (1981).</li><li>Böck, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Romeis Mikroskopische Technik. , 1-697 (1989).</li><li>Suvarna, K. S., Layton, C., Bancroft, J. D., Suvarna, K. S., Layton, C., Bancroft, J. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Bancroft's theory and practice of histological techniques. , 1-654 (2013).</li><li>Knust, J., Ochs, M., Gundersen, H. J., Nyengaard, J. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Stereological+estimates+of+alveolar+number+and+size+and+capillary+length+and+surface+area+in+mice+lungs.">Stereological estimates of alveolar number and size and capillary length and surface area in mice lungs.</a> Anat Rec (Hoboken). 292, 113-122 (2009).</li><li>Nyengaard, J. R., Gundersen, H. J. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sampling+for+stereology+in+lungs.">Sampling for stereology in lungs.</a> Eur Respir Rev. 15, 107-114 (2006).</li><li>Junatas, K. 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著者が明らかに何もありません。

ブタの動物モデルからのバイオバンク サンプル コレクションの生成臓器ボリュームによる冗長な組織サンプルの広い範囲に適しています代表者の再現可能な世代堅牢な技術とプロトコルが必要です。異なる分析方法と薬剤の定量分析のためのサンプル セクションの方向のランダム化のため。本稿で説明する方法は、ブタの臓器や組織のサイズに合わせられるし、これら要求2</s...

大動物モデル1,2,3,4、5、ブタの間いくつかの有利な類似のために医学の豚用ますます一般的なものと人体解剖学と生理学との生成を可能にする確立された分子生物的方法の可用性は、合わせて、豚モデル疾患条件1,4の広い範囲のための遺伝子組換え。
しかし、齧歯動物モデルと比較して、いつでも実験のため提供することができますそれぞれの豚のモデルの動物の数は限られています。これは約 1 年と世代の豚のモデルおよび動物飼育のために必要な金融と時間のかかる努力のブタの世代間隔が原因です。したがって、これらの豚から生成することができますサンプルと同様に、ブタのモデルの個々 の動物、遺伝子組み換え豚モデルおよび/または長期の実験問題 (例えば、晩期合併症の場合は特に、非常に貴重です高齢者2,6,7慢性疾患) について説明します。
後で関連性の高いことが判明して可能性がありますいない研究の初期の実験的設計で予定されていた追加解析のパフォーマンス任意の研究の過程で例えば、以前から生じる個別の質問を発見したアドレスに意外な発見。このような追加実験のための適切なサンプルが利用できない disproportionally 高いコストと時間のかかる支出は、追加の豚や組織サンプルを生成する必要あります。そのような事態に備えてするには、バイオバンクのさまざまな器官、組織、またはバイオ液体、量的そして質的幅広い以降の解析に適した保存バックアップ サンプル集の生成、重要と考えられる2,6,7に近づきます。ブタの動物モデルからの最適の利点を派生する、適切なバイオバンク サンプルの可用性で多臓器レベルで同一のサンプル素材にさまざまな分析方法の広範なスペクトルを実行する独自の可能性を提供しています、非常に同じ個々 の動物、例えば、さまざまなワーキング ・ グループの科学者にサンプルの分布によって組織された研究ネットワーク2,6,7。さらに、biobanking の「将来」のサンプリング戦略研究に必要な動物数削減にも貢献しています。ブタのモデル biobanking の利点は、最近多臓器、臓器を調べる multiomics 研究で実証されているミュンヘン MIDY 豚バイオバンクから試験片を使用して、長期的な糖尿病の遺伝子組み換えブタモデルにおけるクロストーク2。
実行後の分析の結果の解釈可能性と信頼性を確立する一般的に遵守しなければならないバイオバンク サンプルいくつかの必須要件があります。サンプルを再現性をもって発生する必要があります、彼らは十分に代表、すなわちをある必要があります7から採取した組織・臓器の興味の形態学的および分子機能を反映しています。下流解析のタイプの広い範囲に適していること、サンプルする必要があります十分な量での撮影、説明を含む様々 な分析方法の要求 (時間と温度条件を含む) に従って処理されます。cryohistology、パラフィン、プラスチック組織学、免疫組織化学の in situハイブリダイゼーション、微細構造の電子顕微鏡解析、臨床検査診断分析、同様に分子などの病理組織学的解析DNA、RNA、蛋白質、および代謝物の分析。
番号、ボリューム、長さ、表面積の異なる組織構造など個別の定量形態学的パラメーターの広い範囲の評価定量的薬剤分析できるように、ランダム化されたセクションの平面、それぞれの臓器・組織の組織サンプルは、7、8,9,10,11を準備する必要があります。(すなわち、空間参照) から撮影されたサンプルの定量形態学的研究、組織や臓器、器官のコンパートメントの合計量の精密測定は、決定的に重要な7,9,12それぞれの臓器、組織、または有機体内の興味がパラメーターの絶対的な数量を計算します。最終的には、組織切片の作製中に埋め込みに関連する組織収縮効果が決定しアカウント13に。したがって、定量的な薬剤試料の分析、特にアーカイブ (組織サンプル、埋め込まれたティッシュのブロック、標本等を固定) 前の調査から、時々 厳しく制限あるいは不可能12、します。数と利用可能な個々 のサンプルの量が十分な場合、代表的なサンプルを保証する十分なサンプリングのデザインが適用されていない場合、それぞれの臓器・組織の超高速を実行しなかった場合に特にまたは場合の処理、サンプルは、関心の定量形態学的パラメーターの推定と互換性がありません。マニホールドの可能な要因による個別の定量形態学的パラメーターの解析アーカイブ サンプル材料の適合性ははっきりと答えることができない、個々 のケースごとの慎重な評価によります。
したがって、場所、サイズ、数、処理、トリミング方向、およびサンプルの向きが可能性があります以降の解析の結果に影響を与える、これらの要因は非常に重要なは、任意の研究の実験の設計で考慮する必要があります。これらの側面およびブタの解剖学、ブタに適応の動物モデルが確立されている最近の包括的な, 詳細な, 大規模なサンプリング ガイドラインの特徴に関して標準化された、再現性のある堅牢な参照を提供します。、および 50 を超える異なるブタ臓器や組織6,7から冗長、適切に処理された、高品質のサンプルの効率的な生成。
方法論の説明と本稿で示すビデオ チュートリアルを提供様々 な超高速、技術の実用性能の手順詳細、説明でわかりやすくの手順でブタ組織のサンプリングと臓器と異なる下流解析のための組織サンプルの処理。関連組織の三次元収縮寸法の測定を含む注目のテクニックは臓器のボリュームとアルキメデスとカヴァリエーリの9の原則に基づいて密度の定量方法など、病理検査の処理中に別の埋め込みメディア14に埋め込み、実用的な出来高で加重された体系的なランダム サンプリングのアプリケーションに近づくと、サンプリングされた組織検体の処理別以降分析7,8,9,15と世代の適切な指向し、潜在的な薬剤の定量的解析7、8のサンプルを処理 9,10,11。ブタ バイオバンクのプロジェクトで彼らのアプリケーションの横に示されたメソッドは、一般的にすべての臓器・組織の病理組織形態の定量的なプロパティを調べることの研究の適切です。さらに、体系的なランダム サンプリングのデザインが世代の豊富な変化、例えばRna、蛋白質、または代謝産物を検出する分子の解析手法を用いた実験の代表的なサンプルのため特に有益ですさまざまな臓器や組織。
次の段落は、その実用性能がプロトコル」に記載されている間にこれらのメソッドを簡単に紹介を提供します。
臓器ボリュームの定量 器官の重量とボリュームの決定は、いくつかの実験的設定の重要な可能性がありますこれらの要因として実験的に関連する可能性のある変更を調べた関心の要因臓器・組織の総量もよく絶対定量的なパラメーター (例えば、総セル数)、stereologically 推定数値ボリューム密度 (すなわち、ボリューム単位セルの数からの計算に必要な組織) の7,12。コンピューター断層撮影などの複雑な技術的な装置を使用してのテクニックとは別に基本的に器官またはティッシュの絶対量を決定するために一般的に使用される 3 つの実用的な方法があります。器官のボリュームは、「直接体積測定」すなわち、アルキメデスの原理によると、水または完全に浸漬した場合の構造によって転置された生理食塩水の量を測定して決定できます。ただし、対等に大きいブタ器官のこれらのアプローチは非現実的であり、不正確さをしやすい非常に大きい容積測定フラスコを要求するので。便利に、臓器・組織のボリュームはその密度と重量7,12,16、「水没法」7,12 を使用して効率的に判断できます。 から計算できます ,16 (プロトコル手順 1.1)。(1598-1647)「カバリエリの原理」に基づく超高速アプローチを使用しても、臓器・組織ボリュームを推定できます。簡単に言えば、カヴァリエリの原理の状態、地面に平行平面の区分である 2 つのオブジェクトと対応するで 2 つのオブジェクトをセクションのプロファイルをカットする場合、地面からの距離で 2 つのオブジェクト、同じエリアがあります。同じボリュームを持っています。したがって、並列、等しく遠く離れている断面平面と断面平面間の距離で、プロファイル断面の製品として任意形状オブジェクトのボリュームを推定できます。これは次の類推で理解できる: 同じ硬貨の同じ数から成る 2 つのスタックは、サイドバイ サイド、コイン整然としたコイン スタック等の円筒形の降伏、互いの上に積み上げ、1 つのスタックの配置検討します。オフセンターの硬貨のスタックには、コイン (図 3 a) が配置されます。両方のコイン スタックの形状が異なる、ボリュームが同じで、両方のスタックの対応するレベルでコインの区域以来 (すなわち、並列セクションのプロフィールの部分はから同じ距離で両方のコイン スタックを介してカット、地面) と同じです。ブタの臓器や組織は、カヴァリエリの原理7,12,15を使用してのボリュームの推定、1.2 の手順で説明します。
組織学的埋め込みに関連する組織の収縮の程度の決定 組織切片で測定するいくつかの定量的形態学的パラメーターの解析では、ティッシュの組織学のための処理中に発生した埋め込みに関連する組織収縮効果決定し、考慮しています。埋め込みに関連する組織の収縮の範囲変数、組織、その処理と埋め込み中8,13,17,18,19に依存します。一般的に、埋め込み関連組織サンプル (すなわち、ほとんど収縮) の容積の変化領域、および、したがって、すべての 3次元パラメーター推定定量化薬剤分析8 に影響を与えるすべての 3 つの次元で.基本的には、手順 1.3 に示すように埋め込みに関連する組織の収縮は、線形組織収縮因子 (fS) として表現の程度を推定できます。(収縮に敏感な) 定量的形態パラメーター14の補正のために使用されます。
出来高で加重された臓器・組織の体系的なランダム サンプリング ブタ臓器試料のバイオバンク コレクションの構築、手順 2 で説明したように出来高で加重された体系的なランダム サンプリングの手法が代表者の世代の実用的な時間の節約と効率的なテクニックをあることを証明します。多目的組織サンプル7,8,9,15です。
薬剤の定量解析の等方性均一ランダム セクションおよび均一ランダム垂直セクションの生成 バイオバンク組織サンプルは、適切に準備された標本なしできなかったパラメーターの最大値の推定のための異なる定量的解析方法の広い範囲に適している必要があります。「等方性 (独立) 均一ランダム (フェース) セクション「8,9を使用して、ほぼすべての定量的な主パラメーターを決定できます。フェース セクションで組織サンプルの断面平面の 3次元配向がランダムします。これは、断面平面の位置を基準にして組織サンプルの位置のランダム化が適用されると"Isector"法11 (プロトコル手順 3.1)、相対的な断面平面の方向のランダム化に実現、組織サンプルは、"Orientator"法10 (プロトコル手順 3.2) のように。皮膚または粘膜標本自然存在または定義され適切に識別垂直軸、「垂直一様ランダム (VUR) セクション」の準備を表示するなどの組織サンプルの (プロトコル手順 3.3) は厳密の平面内断面の縦軸は有利な8,20です。フェース/VUR サンプリングの理論的基礎と潜在的な下流定量化薬剤分析の包括的な議論の完全な談話、興味を持った読者定量的ステレオロジー人生の教科書に呼びます科学8,9。

<table>
	<tbody>
		<tr>
			<td>Agar</td>
			<td>Carl Roth GmbH, Germany</td>
			<td>Agar (powder), Cat.: 5210.3</td>
			<td>Dissolve approximately 1 g of agar in 10 ml cold water in a glass or plastic beaker, heat in microwave-oven at 700 W, boil the solution twice with rigorous stirring. Cast into mold while still warm and let solidify. Caution: While handling with hot liquid agar, wear protective goggles and gloves.</td>
		</tr>
		<tr>
			<td>Caliper</td>
			<td>Hornbach Baumarkt GmbH, Bornheim, Germany</td>
			<td>Schieblehre Chrom/Vernickelt 120 mm Cat.: 3664902</td>
			<td>Any kind of caliper (mechanical or electronic) will do as well.</td>
		</tr>
		<tr>
			<td>Casting molds (metal)</td>
			<td>Engelbrecht Medizin &#38; Labortechnik, Ederm&#252;nde, Germany</td>
			<td>Einbettsch&#228;lchen aus Edelstahl, 14 x 24 x 5 mm, Cat.: 14302b</td>
			<td>Any other kind of metal casting mold used for paraffin-embedding will do as well.</td>
		</tr>
		<tr>
			<td>Copy templates of cross grids (5mm - 6 cm)</td>
			<td>n.a.</td>
			<td>n.a.</td>
			<td>Copy templates of cross grids (5mm - 6 cm) are provided in the supplemental data file of Albl et al.&#160; Toxicol Pathol. 44, 414-420, doi: 10.1177/0192623316631023 (2016)</td>
		</tr>
		<tr>
			<td>Copy templates of equiangular and cosine-weighted circles</td>
			<td>n.a.</td>
			<td>n.a.</td>
			<td>Copy templates of equiangular and cosine-weighted circles are provided in Nyengaard &#38; Gundersen. Eur Respir Rev. 15, 107-114, doi: 10.1183/09059180.00010101 (2006) and in Gundersen et al. Stereological Principles and Sampling Procedures for Toxicologic Pathologists. In: Haschek and Rousseaux&#180;s Handbook of Toxicologic Pathology. 3rd ed, 215-286, ISBN: 9780124157590 (2013).</td>
		</tr>
		<tr>
			<td>Foldback clamps (YIHAI binder clips, 15 mm and 19 mm)</td>
			<td>Ningbo Tianhong Stationery Co ltd., China</td>
			<td>Y10006 and Y10005</td>
			<td>Any other type of standard office foldback clamps will do as well.</td>
		</tr>
		<tr>
			<td>Forceps (anatomical)</td>
			<td>NeoLab Migge GmbH, Heidelberg, Germany</td>
			<td>neoLab Standard -Pinzette 130 mm, anatomisch, rund, Cat.: 1-1811</td>
			<td>Any type of anatomical forceps will do.</td>
		</tr>
		<tr>
			<td>Formaldehyde-solution 4%</td>
			<td>SAV-Liquid Produktion GmbH, Flintsbach, Germany</td>
			<td>Formaldehyd 37/40 %, Cat.: 1000411525005</td>
			<td>Dilute to 4% from concentrated solution. Buffer to neutral pH. Wear appropriate eye-, hand- and respiratory protection. Process tissue samples fixed in formaldehyde solution under an exhaust hood and wear protective goggles and laboratory gloves.</td>
		</tr>
		<tr>
			<td>Graph paper (for calibration)</td>
			<td>B&#252;romarkt B&#246;ttcher AG, Jena, Germany. www.bueromarkt-ag.de</td>
			<td>Penig Millimeterpapier A4, Cat.: 2514</td>
			<td>Any type of graph paper (scaled in millimeter) will do.</td>
		</tr>
		<tr>
			<td>Laboratory beakers (5ml, 10 ml, 50 ml, 100 ml)</td>
			<td>NeoLab Migge GmbH, Heidelberg, Germany</td>
			<td>Becherglas SIMAX&#174; , niedrige Form, Borosilikatglas 3.3 Cat.: E-1031, E-1032, E-1035, E-1036</td>
			<td>Any kind of glass- or plastic beakers of 5 &#8211; 100 ml volume will do.</td>
		</tr>
		<tr>
			<td>Laboratory scale(s)</td>
			<td>Mettler Toledo GmbH, Gie&#223;en, Germany</td>
			<td>PM6000</td>
			<td rowspan="2">Any standard laboratory scales with measuring ranges between 0.1 mg to approximately 20 g, respectively between 100 mg to approximately 500 g will do</td>
		</tr>
		<tr>
			<td></td>
			<td>Sartorius AG, G&#246;ttingen, Germany</td>
			<td>BP61S</td>
		</tr>
		<tr>
			<td>Microtome blades</td>
			<td>Engelbrecht Medizin &#38; Labortechnik, Ederm&#252;nde, Germany</td>
			<td>FEATHER Microtome blasdes S35, Cat.:14700</td>
			<td>Any kind of single-use microtome blades will do.</td>
		</tr>
		<tr>
			<td rowspan="2">Morphometry/planimetry software/system</td>
			<td>National Institute of Health (NIH)</td>
			<td>ImageJ</td>
			<td>Download from https://www. imagej.nih.gov/ij/ (1997).</td>
		</tr>
		<tr>
			<td>Zeiss-Kontron, Eching, Germany</td>
			<td>VideoplanTM image analysis system</td>
			<td>Out of stock</td>
		</tr>
		<tr>
			<td>Photo camera</td>
			<td>Nikon</td>
			<td>D40</td>
			<td>Any kind of digital photocamera that can be mounted to a tripod&#160; will do.</td>
		</tr>
		<tr>
			<td>Plastic transparencies</td>
			<td>Avery Zweckform GmbH, Oberlaindern, Germany</td>
			<td>Laser Overhead-Folie DINA4 Cat.:&#160; 3562</td>
			<td>Any (laser)-printable plastic transparency will do.</td>
		</tr>
		<tr>
			<td>Random number tables</td>
			<td>n.a.</td>
			<td>n.a.</td>
			<td>Random number tables can conveniently be generated (with defined numbers of random numbers and within defined intervals), using random number generators, such as: https://www.random.org/</td>
		</tr>
		<tr>
			<td>Razor blades</td>
			<td>Plano GmbH, Wetzlar, Germany</td>
			<td>T5016</td>
			<td>Any kind of razor blades will do.</td>
		</tr>
		<tr>
			<td>Ruler</td>
			<td>B&#252;romarkt B&#246;ttcher AG, Jena, Germany. www.bueromarkt-ag.de</td>
			<td>Office-Point Lineal 30 cm, Kunststoff, transparent, Cat.: ln30</td>
			<td>Any kind of cm-mm-scaled ruler will do as well.</td>
		</tr>
		<tr>
			<td>Saline (0.9%)</td>
			<td>Carl Roth GmbH, Germany</td>
			<td>Natriumchlorid, &#62;99% Cat.: 0601.1</td>
			<td>To prepare 0.9% saline, dissolve 9 g NaCl in 1000 ml of distilled water at 20&#176;C.</td>
		</tr>
		<tr>
			<td>Scalpel blades</td>
			<td>Aesculap AG &#38; Co KG, Tuttlingen, Germany</td>
			<td>BRAUN Surgical blades N&#176;22</td>
			<td>Any kind of scalpel blades will do.</td>
		</tr>
		<tr>
			<td>Scanner</td>
			<td>Hewlett-Packard</td>
			<td>hp scanjet 7400c</td>
			<td>Any type of standard office scanner capable of scanning with resolutions from 150-600 dpi will do.</td>
		</tr>
		<tr>
			<td>Slicing devices</td>
			<td>n.a.</td>
			<td>n.a.</td>
			<td>Examples forself constructed slicing devices can be found in Knust, et al. Anatomical record. 292, 113-122, doi: 10.1002/ar.20747 (2009) and in the supplemental data file of Albl et al.&#160; Toxicol Pathol. 44, 414-420, doi: 10.1177/0192623316631023 (2016).</td>
		</tr>
		<tr>
			<td>Spherical casting molds (e.g., in 25.5 mm diameter)</td>
			<td>Pralinen-Zutaten.de, Windach, Germany</td>
			<td>Pralinen-Hohlkugeln Vollmilch, 25.5 mm</td>
			<td>Spherical casting molds can as well be be self-constructed, or obtained from other confectioner suppliers (for for pralines). The casting molds indicated here are actually the package/wrapping of hollow pralines bodies (first eat the pralines and then use the package for generation of i-sector sections)</td>
		</tr>
		<tr>
			<td>Thin wire</td>
			<td>Basteln &#38; Hobby Schobes, Stra&#223;furth, Germany. www,bastel-welt.de</td>
			<td>Messingdraht (0.3 mm) Cat.: 216464742</td>
			<td>Any other kind of thin wire will also do.</td>
		</tr>
		<tr>
			<td>Tissue paper</td>
			<td>NeoLab Migge GmbH, Heidelberg, Germany</td>
			<td>Declcate Task Wipes-White, Cat.: 1-5305</td>
			<td>Any other kind of laboratory tissue paper will do as well.</td>
		</tr>
		<tr>
			<td>Waterproof pen</td>
			<td>Staedler Mars GmbH &#38; Co KG, N&#252;rnberg, Grmany</td>
			<td>Lumocolor permanent 313, 0.4 mm, S, black, Cat.: 313-2</td>
			<td>Any other kind of waterproof pen will do as well.</td>
		</tr>
	</tbody>
</table>

sampling strategies and processing of biobank tissue samples from porcine biomedical models

医学トランスレーショナルリサーチのブタのモデルはより人気となって着実にいます。モデル、およびこれらのモデルの使用可能な動物の多くの場合限られた数の遺伝子組み換え豚を中心に、個々 の動物の高い値を考慮した、適切に処理された組織サンプル集 (バイオバンク) の確立に最適、以降の解析方法、サンプリングの時点で指定されていない解析などの広範なスペクトルを表すモデルの直線移動の値をフル活用する意味のあるアプローチ。ブタの解剖学の特性に関して包括的なガイドラインは標準化された世代代表、異なるブタの臓器や組織からの質の高いサンプルの最近確立されています。これらのガイドラインは、結果および比較可能性さまざまな研究と研究者間の再現性の重要な前提条件です。臓器重量とボリューム、生成される定量サンプリング場所および組織サンプルの番号と同様に自分の向き、サイズ、処理およびトリム方向などの基本的なデータの記録、関連する要因一般化と分子、定性的手法と定量的形態解析用の標本の有用性を決定します。ここでは、代表の世代の最も重要な技術の説明、実用的なステップバイ ステップ サンプル、ブタの組織から多目的バイオバンク検体が提示されます。ここで説明した方法があります決定臓器のボリュームと密度の実質臓器組織収縮の範囲のポイントを数える測定による出来高で加重された体系的なランダム サンプリング プロシージャのアプリケーションサンプル、および垂直な制服と"Orientator"と"Isector"メソッドが生成された等方性一様ランダム (フェース) セクションなど、定量的な薬剤分析ランダム配向試料の世代の組織学的埋め込みに関連ランダム (VUR) セクション。

水没組織・臓器密度の測定法
図 12A-Bは、1.1 (図 1図 2) のステップで説明されている水没を用いたブタ腎臓のボリュームと密度の代表決定を示しています。追加豚の臓器や組織の密度測定の代表的な結果は、表 2に掲載されています。...

Watch this Scientific Journal Video about Sampling Strategies and Processing of Biobank Tissue Samples from Porcine Biomedical Models at JoVE.com

Sampling Strategies and Processing of Biobank Tissue Samples from Porcine Biomedical Models

サンプリング戦略とブタの生物医学的モデルからバイオバンク組織サンプルの処理

バイオバンク プロジェクトの下流解析の広範なスペクトルのブタの動物モデルの代表的な組織サンプルの生成法の性能と実用を実証、超高速を含む体系的なランダム サンプリング定性・定量形態学上および分子分析型の組織サンプルの差分処理。

In translational medical research, porcine models have steadily become more popular. Considering the high value of individual animals, particularly of ...

sampling-strategies-processing-biobank-tissue-samples-from-porcine

Research

JoVE Journal

Medicine

15.5K ビュー.  LMU Munich. バイオバンク プロジェクトの下流解析の広範なスペクトルのブタの動物モデルの代表的な組織サンプルの生成法の性能と実用を実証、超高速を含む体系的なランダム サンプリング定性・定量形態学上および分子分析型の組織サンプルの差分処理。

ビデオ: Sampling Strategies and Processing of Biobank Tissue Samples from Porcine Biomedical Models

Generation of Alginate Microspheres for Biomedical Applications

Alginate-based materials have received considerable attention for biomedical applications because of their hydrophilic nature, biocompatibility, and physical architecture. Applications include cell encapsulation, drug delivery, stem cell culture, and tissue engineering scaffolds. In fact, clinical trials are currently being performed in which islets are encapsulated in PLO coated alginate microbeads as a treatment of type I diabetes. However, large numbers of islets are required for efficacy due to poor survival following transplantation. The ability to locally stimulate microvascular network formation around the encapsulated cells may increase their viability through improved transport of oxygen, glucose and other vital nutrients. Fibroblast growth factor-1 (FGF-1) is a naturally occurring growth factor that is able to stimulate blood vessel formation and improve oxygen levels in ischemic tissues. The efficacy of FGF-1 is enhanced when it is delivered in a sustained fashion rather than a single large-bolus administration. The local long-term release of growth factors from islet encapsulation systems could stimulate the growth of blood vessels directly towards the transplanted cells, potentially improving functional graft outcomes. In this article, we outline procedures for the preparation of alginate microspheres for use in biomedical applications. In addition, we describe a method we developed for generating multilayered alginate microbeads. Cells can be encapsulated in the inner alginate core, and angiogenic proteins in the outer alginate layer. The release of proteins from this outer layer would stimulate the formation of local microvascular networks directly towards the transplanted islets.

In the following sections, we outline procedures for the preparation of alginate microspheres for use in biomedical applications. We specifically illustrate a technique for creating multilayered alginate microspheres for the dual purpose of cell and protein encapsulation as a potential treatment for type 1 diabetes.

バイオメディカル応用のためのアルギン酸ミクロスフェアの生成

Alginate-based materials have received considerable attention for biomedical applications because of their hydrophilic nature, biocompatibility, and ...

医学

How to Measure Cortical Folding from MR Images: a Step-by-Step Tutorial to Compute Local Gyrification Index

Cortical folding (gyrification) is determined during the first months of life, so that adverse events occurring during this period leave traces that will be identifiable at any age. As recently reviewed by Mangin and colleagues2, several methods exist to quantify different characteristics of gyrification. For instance, sulcal morphometry can be used to measure shape descriptors such as the depth, length or indices of inter-hemispheric asymmetry3. These geometrical properties have the advantage of being easy to interpret. However, sulcal morphometry tightly relies on the accurate identification of a given set of sulci and hence provides a fragmented description of gyrification. A more fine-grained quantification of gyrification can be achieved with curvature-based measurements, where smoothed absolute mean curvature is typically computed at thousands of points over the cortical surface4. The curvature is however not straightforward to comprehend, as it remains unclear if there is any direct relationship between the curvedness and a biologically meaningful correlate such as cortical volume or surface. To address the diverse issues raised by the measurement of cortical folding, we previously developed an algorithm to quantify local gyrification with an exquisite spatial resolution and of simple interpretation. Our method is inspired of the Gyrification Index5, a method originally used in comparative neuroanatomy to evaluate the cortical folding differences across species. In our implementation, which we name local Gyrification Index (lGI1), we measure the amount of cortex buried within the sulcal folds as compared with the amount of visible cortex in circular regions of interest. Given that the cortex grows primarily through radial expansion6, our method was specifically designed to identify early defects of cortical development.
In this article, we detail the computation of local Gyrification Index, which is now freely distributed as a part of the FreeSurfer Software (<a href="http://surfer.nmr.mgh.harvard.edu/">http://surfer.nmr.mgh.harvard.edu/</a>, Martinos Center for Biomedical Imaging, Massachusetts General Hospital). FreeSurfer provides a set of automated reconstruction tools of the brain's cortical surface from structural MRI data. The cortical surface extracted in the native space of the images with sub-millimeter accuracy is then further used for the creation of an outer surface, which will serve as a basis for the lGI calculation. A circular region of interest is then delineated on the outer surface, and its corresponding region of interest on the cortical surface is identified using a matching algorithm as described in our validation study1. This process is repeatedly iterated with largely overlapping regions of interest, resulting in cortical maps of gyrification for subsequent statistical comparisons (Fig. 1). Of note, another measurement of local gyrification with a similar inspiration was proposed by Toro and colleagues7, where the folding index at each point is computed as the ratio of the cortical area contained in a sphere divided by the area of a disc with the same radius. The two implementations differ in that the one by Toro et al. is based on Euclidian distances and thus considers discontinuous patches of cortical area, whereas ours uses a strict geodesic algorithm and include only the continuous patch of cortical area opening at the brain surface in a circular region of interest.

Measuring gyrification (cortical folding) at any age represents a window into early brain development. Hence, we previously developed an algorithm to measure local gyrification at thousands of points over the hemisphere1. In this paper, we detail the computation of this local gyrification index.

ローカルGyrificationのインデックスを計算するステップバイステップのチュートリアル：MR画像からの皮質折りたたみを測定する方法

Cortical folding (gyrification) is determined during the first months of life, so that adverse events occurring during this period leave traces that will ...

Processing of Primary Brain Tumor Tissue for Stem Cell Assays and Flow Sorting

Brain tumors are typically comprised of morphologically diverse cells that express a variety of neural lineage markers. Only a relatively small fraction of cells in the tumor with stem cell properties, termed brain tumor initiating cells (BTICs), possess an ability to differentiate along multiple lineages, self-renew, and initiate tumors in vivo. We applied culture conditions originally used for normal neural stem cells (NSCs) to a variety of human brain tumors and found that this culture method specifically selects for stem-like populations. Serum-free medium (NSC) allows for the maintenance of an undifferentiated stem cell state, and the addition of bFGF and EGF allows for the proliferation of multi-potent, self-renewing, and expandable tumorspheres.
To further characterize each tumor's BTIC population, we evaluate cell surface markers by flow cytometry. We may also sort populations of interest for more specific characterization. Self-renewal assays are performed on single BTICs sorted into 96 well plates; the formation of tumorspheres following incubation at 37 &deg;C indicates the presence of a stem or progenitor cell. Multiple cell numbers of a particular population can also be sorted in different wells for limiting dilution analysis, to analyze self-renewal capacity. We can also study differential gene expression within a particular cell population by using single cell RT-PCR.
The following protocols describe our procedures for the dissociation and culturing of primary human samples to enrich for BTIC populations, as well as the dissociation of tumorspheres. Also included are protocols for staining for flow cytometry analysis or sorting, self-renewal assays, and single cell RT-PCR.

The identification of brain tumor initiating cells (BTICs), the rare cells within a heterogeneous tumor possessing stem cell properties, provides new insights into human brain tumor pathogenesis. We have refined specific culture conditions to enrich for BTICs, and we routinely use flow cytometry to further enrich these populations. Self-renewal assays and transcript analysis by single cell RT-PCR can subsequently be performed on these isolated cells.

ソート幹細胞アッセイおよびフローの原発性脳腫瘍組織の処理

Brain tumors are typically comprised of morphologically diverse cells that express a variety of neural lineage markers. Only a relatively small fraction ...

Isolation and Functional Characterization of Human Ventricular Cardiomyocytes from Fresh Surgical Samples

Cardiomyocytes from diseased hearts are subjected to complex remodeling processes involving changes in cell structure, excitation contraction coupling and membrane ion currents. Those changes are likely&#160;to be responsible for the increased arrhythmogenic risk and the contractile alterations leading to systolic and diastolic dysfunction in cardiac patients. However, most information on the alterations of myocyte function in cardiac diseases has come from animal models.
Here we describe and validate a protocol to isolate viable myocytes from small surgical samples of ventricular myocardium from patients undergoing cardiac surgery operations. The protocol is described in detail. Electrophysiological and intracellular calcium measurements are reported to demonstrate the feasibility of a number of single cell measurements in human ventricular cardiomyocytes obtained with this method.
The protocol reported here can be useful for future investigations of the cellular and molecular basis of functional alterations of the human heart in the presence of different cardiac diseases. Further, this method can be used to identify novel therapeutic targets at cellular level and to test the effectiveness of new compounds on human cardiomyocytes, with direct translational value.

Current knowledge on the cellular basis of cardiac diseases mostly relies on studies on animal models. Here we describe and validate a novel method to obtain single viable cardiomyocytes from small surgical samples of human ventricular myocardium. Human ventricular myocytes can be used for electrophysiological studies and drug testing.

離と新鮮な外科的試料からのヒト心室の心筋細胞の機能解析

Cardiomyocytes from diseased hearts are subjected to complex remodeling processes involving changes in cell structure, excitation contraction coupling and ...

The Bovine Lung in Biomedical Research: Visually Guided Bronchoscopy, Intrabronchial Inoculation and In Vivo Sampling Techniques

There is an ongoing search for alternative animal models in research of respiratory medicine. Depending on the goal of the research, large animals as models of pulmonary disease often resemble the situation of the human lung much better than mice do. Working with large animals also offers the opportunity to sample the same animal repeatedly over a certain course of time, which allows long-term studies without sacrificing the animals.
The aim was to establish in vivo sampling methods for the use in a bovine model of a respiratory Chlamydia psittaci infection. Sampling should be performed at various time points in each animal during the study, and the samples should be suitable to study the host response, as well as the pathogen under experimental conditions.
Bronchoscopy is a valuable diagnostic tool in human and veterinary medicine. It is a safe and minimally invasive procedure. This article describes the intrabronchial inoculation of calves as well as sampling methods for the lower respiratory tract. Videoendoscopic, intrabronchial inoculation leads to very consistent clinical and pathological findings in all inoculated animals and is, therefore, well-suited for use in models of infectious lung disease. The sampling methods described are bronchoalveolar lavage, bronchial brushing and transbronchial lung biopsy. All of these are valuable diagnostic tools in human medicine and could be adapted for experimental purposes to calves aged 6-8 weeks. The samples obtained were suitable for both pathogen detection and characterization of the severity of lung inflammation in the host.

The Bovine Lung in Biomedical Research: Visually Guided Bronchoscopy, Intrabronchial Inoculation and In Vivo Sampling Techniques

This article describes bronchoscopic techniques in the bovine lung under experimental conditions, i.e. bronchoscopically guided inoculation, bronchoalveolar lavage, bronchial brushing, and transbronchial lung biopsy.

生物医学研究における牛の肺：視覚ガイド付き気管支鏡検査、気管支内接種および<em&gt;インビボ</em&gt;サンプリングテクニック

There is an ongoing search for alternative animal models in research of respiratory medicine. Depending on the goal of the research, large animals as ...

Unbiased Deep Sequencing of RNA Viruses from Clinical Samples

Here we outline a next-generation RNA sequencing protocol that enables de novo assemblies and intra-host variant calls of viral genomes collected from clinical and biological sources. The method is unbiased and universal; it uses random primers for cDNA synthesis and requires no prior knowledge of the viral sequence content. Before library construction, selective RNase H-based digestion is used to deplete unwanted RNA &#8212; including poly(rA) carrier and ribosomal RNA &#8212; from the viral RNA sample. Selective depletion improves both the data quality and the number of unique reads in viral RNA sequencing libraries. Moreover, a transposase-based &#39;tagmentation&#39; step is used in the protocol as it reduces overall library construction time. The protocol has enabled rapid deep sequencing of over 600 Lassa and Ebola virus samples-including collections from both blood and tissue isolates-and is broadly applicable to other microbial genomics studies.

This protocol describes a rapid and broadly applicable method for unbiased RNA-sequencing of viral samples from human clinical isolates.

臨床サンプルからのRNAウイルスの公平なディープシーケンシング

Here we outline a next-generation RNA sequencing protocol that enables de novo assemblies and intra-host variant calls of viral genomes collected from ...

Absorption of Nasal and Bronchial Fluids: Precision Sampling of the Human Respiratory Mucosa and Laboratory Processing of Samples

The methods of nasal absorption (NA) and bronchial absorption (BA) use synthetic absorptive matrices (SAM) to absorb the mucosal lining fluid (MLF) of the human respiratory tract. NA is a non-invasive technique which absorbs fluid from the inferior turbinate, and causes minimal discomfort. NA has yielded reproducible results with the ability to frequently repeat sampling of the upper airway. 
By comparison, alternative methods of sampling the respiratory mucosa, such as nasopharyngeal aspiration (NPA) and conventional swabbing, are more invasive and may result in greater data variability. Other methods have limitations, for instance, biopsies and bronchial procedures are invasive, sputum contains many dead and dying cells and requires liquefaction, exhaled breath condensate (EBC) contains water and saliva, and lavage samples are dilute and variable.
BA can be performed through the working channel of a bronchoscope in clinic. Sampling is well tolerated and can be conducted at multiple sites in the airway. BA results in MLF samples being less dilute than bronchoalveolar lavage (BAL) samples.
This article demonstrates the techniques of NA and BA, as well as the laboratory processing of the resulting samples, which can be tailored to the desired downstream biomarker being measured. These absorption techniques are useful alternatives to the conventional sampling techniques used in clinical respiratory research.

This manuscript describes the use of nasal and bronchial absorption techniques, specifically using synthetic absorptive matrices (SAM) to sample the mucosal lining fluid (MLF) of the upper and lower airway. These methods provide better standardization and tolerability than existing respiratory sampling techniques.

鼻粘膜および気管支の流体の吸収: 人間の呼吸器粘膜とサンプル室加工高精度サンプリング

The methods of nasal absorption (NA) and bronchial absorption (BA) use synthetic absorptive matrices (SAM) to absorb the mucosal lining fluid (MLF) of the ...

Improvement of a Closed Chest Porcine Myocardial Infarction Model by Standardization of Tissue and Blood Sampling Procedures

Myocardial ischemia reperfusion (I/R) injury contributes to almost half of the necrotic area after myocardial infarction. To date there is no approved drug to prevent or reduce myocardial I/R injury. The study and understanding of the pathophysiological mechanisms of myocardial I/R injury is essential to develop successful treatments. Large animal experiments are an important step in translational methods. The porcine model of acute myocardial infarction has been established and described by ourselves and others. We aimed to further improve the value of the model by focusing in detail on the sampling techniques for use in future experiments. Furthermore, we emphasize small but important steps that can affect the quality of the final results. To mimic the clinical situation of myocardial I/R injury, a percutaneous coronary intervention (PCI) catheter was inserted into the left anterior descending coronary artery (LAD) of an anesthetized pig. &#176;&#176;&#176;This model mimics acute myocardial infarction and PCI treatment in humans with the possibility of accurately determining the area at risk as well as the necrotic- and viable ischemic tissue. Here the model was used to investigate the effect of a bicyclic peptide inhibitor of FXIIa. The model can also be modified to allow longer reperfusion times to study later effects of myocardial infarction.

Here we demonstrate a protocol to standardize sampling procedures of an established porcine model of acute myocardial infarction in order to increase its translational value in the understanding of the pathophysiology of myocardial ischemia/reperfusion injury and to test novel drug candidates.

組織と血液サンプリング手順の標準化による閉胸ブタ心筋梗塞モデルの改善

Myocardial ischemia reperfusion (I/R) injury contributes to almost half of the necrotic area after myocardial infarction. To date there is no approved ...

Human Subcutaneous Adipose Tissue Sampling Using a Mini-Liposuction Technique

Studies on adipose tissue are useful in understanding metabolic and other conditions. Human subcutaneous adipose tissue is accessible. With appropriate training and strict adherence to aseptic technique, subcutaneous adipose samples can be safely and efficiently obtained in a non-clinical setting by researchers. Following the administration of local anesthetic lateral to the umbilicus, a 14 G needle attached to a 5 or 10 mL syringe is inserted through the skin into the subcutaneous tissue. Under suction, the syringe is moved in a reciprocating, slicing motion to isolate fragments of adipose tissue. Withdrawing the plunger is enough to ensure that adipose tissue fragments are aspirated through the needle into the syringe. A single biopsy can collect about 200 mg of tissue. This biopsy technique is very safe for both participants and research staff. Following the biopsy, participants can resume most everyday activities, although they should avoid swimming and overly strenuous activities for 48 h to avoid excessive bleeding. Participants can safely undergo 2 biopsies within a single day, meaning that the technique can be applied in before-after acute intervention studies.

The manuscript and associated video demonstrate a percutaneous biopsy technique to obtain samples of subcutaneous adipose tissue from areas surrounding the umbilicus. This method is a low-risk and efficient way to investigate a range of parameters (e.g., gene or protein expression, enzyme activity, lipid content) within adipose tissue.

ミニ脂肪吸引法を用いたヒト皮下脂肪組織サンプリング

Studies on adipose tissue are useful in understanding metabolic and other conditions. Human subcutaneous adipose tissue is accessible. With appropriate ...

Repetitive Blood Sampling from the Subclavian Vein of Conscious Rat

Rats are widely used in pharmacokinetics (PK) and toxicokinetic (TK) studies that need to collect a certain amount of blood at specific time points to detect drug exposure. The rat blood sampling method determines the quality of the plasma and further affects the precision of the test results. The subclavian vein blood collection method described in this protocol collects blood samples repeatedly in the conscious state of animals to meet the needs of PK and TK tests. The skills of restraint handling and appropriate procedure of needle incision ensure the success rate of blood collection. It is easy to operate while ensuring the quality of plasma and at the same time catering to animal welfare. However, this method requires skilled operation, and an improper one may cause animal weakness, pain, lameness, and even mortality. The current method has been used in the testing facility for a 4-week oral toxicity study in Sprague Dawley (SD) rats with TK. The maximum amount of blood collected within 24 h did not exceed 20% of the animal's total blood. The animals' body weight was more than 200 g for males and females. The data showed that the animals' bodyweight increased steadily every week, and the clinical observation was normal after repetitive sample collection.

The present protocol describes a simple and efficient method for collecting blood from the subclavian vein in rats. It enables rapid, timely, and easily identifiable sampling without anesthesia and obtains high-quality blood through repetitive sample collection.

意識性ラットの鎖骨下静脈からの反復採血

Rats are widely used in pharmacokinetics (PK) and toxicokinetic (TK) studies that need to collect a certain amount of blood at specific time points to ...