An accessible and high-throughput strategy of continuously monitoring apoptosis by fluorescent detection of caspase activation.
An accessible and high-throughput strategy of continuously monitoring apoptosis by fluorescent detection of caspase activation.
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通过荧光检测caspase激活来连续监测凋亡的一种可访问且高通量的策略。
DOI：
10.1016/j.ab.2018.10.022
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发表时间：
2019-01-01
影响因子：
2.9
通讯作者：
Finkelstein JN
中科院分区：
文献类型：
--
作者：
Hanson KM;Finkelstein JN
我们提出了一种实时、高通量和成本效益高的体外检测细胞凋亡的方法，该方法使用先前开发的一种通过荧光检测caspase激活的试剂。目前评估细胞凋亡的方法未能考虑到时间维度，因此仅限于每个样本产生的数据。这种试剂可以实时检测细胞凋亡，但到目前为止，这种试剂仅限于昂贵的自动检测系统。在这里，我们描述了Essen Bioscience IncuCyte®Caspase-3/7试剂使用多模式微型平板阅读器进行的细胞凋亡检测，这是生物实验室中的一种常见仪器，可以在自动化系统之前使用，也可以替代自动化系统。根据建立的自动化系统对该改良的微板阅读器的细胞凋亡检测进行了验证，并显示出检测到很强的剂量-反应关系(自动化系统R2=0.9968，微板阅读器R2=0.9924)。我们还提出了一种快速、可靠的方法，通过在微板上进行Hoechst 33342核染色来定量细胞密度(Hoechst信号与细胞密度之间的R2=0.8812)。我们断言，时间维度不应被忽视，并且由于启动成本低和对细胞凋亡的实时准确检测，这里提出的方法对于许多研究人员来说是一种可行的策略。
We present a real-time, high-throughput, and cost-effective method of detecting apoptosis in vitro using a previously developed reagent that detects caspase activation by fluorescence. Current methods of assessing apoptosis fail to account for the dimension of time, and thus are limited in data yielded per sample. This reagent allows real-time detection of apoptosis, but until now has been restricted to a costly automated detection system. Here, we describe apoptosis detection with the Essen Bioscience IncuCyte® Caspase-3/7 Reagent using a multimode microplate reader, a common instrument in biological laboratories, which may be used prior to or in lieu of the automated system. This modified microplate reader apoptosis assay was validated against the established automated system, and was shown to detect a strong dose-response relationship (automated system r2 = 0.9968, microplate reader r2 = 0.9924). We also propose a quick and reliable method of quantifying cell density by Hoechst 33342 nuclear staining in microplates (r2 = 0.8812 between Hoechst signal and cell density). We assert that the dimension of time should not be overlooked, and that the method presented here is an accessible strategy for many researchers due to low startup cost and precise detection of apoptosis in real time.
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影响因子：
23.9
作者：
Bentzinger, C. Florian;Wang, Yu Xin;von Maltzahn, Julia;Soleimani, Vahab D.;Yin, Hang;Rudnicki, Michael A.;Rudnicki, A.
通讯作者：
Rudnicki, A.
影响因子：
5.6
作者：
Hwang HS;Kim HA
通讯作者：
Kim HA
影响因子：
12.4
作者：
Galluzzi L;Vitale I;Aaronson SA;Abrams JM;Adam D;Agostinis P;Alnemri ES;Altucci L;Amelio I;Andrews DW;Annicchiarico-Petruzzelli M;Antonov AV;Arama E;Baehrecke EH;Barlev NA;Bazan NG;Bernassola F;Bertrand MJM;Bianchi K;Blagosklonny MV;Blomgren K;Borner C;Boya P;Brenner C;Campanella M;Candi E;Carmona-Gutierrez D;Cecconi F;Chan FK;Chandel NS;Cheng EH;Chipuk JE;Cidlowski JA;Ciechanover A;Cohen GM;Conrad M;Cubillos-Ruiz JR;Czabotar PE;D'Angiolella V;Dawson TM;Dawson VL;De Laurenzi V;De Maria R;Debatin KM;DeBerardinis RJ;Deshmukh M;Di Daniele N;Di Virgilio F;Dixit VM;Dixon SJ;Duckett CS;Dynlacht BD;El-Deiry WS;Elrod JW;Fimia GM;Fulda S;García-Sáez AJ;Garg AD;Garrido C;Gavathiotis E;Golstein P;Gottlieb E;Green DR;Greene LA;Gronemeyer H;Gross A;Hajnoczky G;Hardwick JM;Harris IS;Hengartner MO;Hetz C;Ichijo H;Jäättelä M;Joseph B;Jost PJ;Juin PP;Kaiser WJ;Karin M;Kaufmann T;Kepp O;Kimchi A;Kitsis RN;Klionsky DJ;Knight RA;Kumar S;Lee SW;Lemasters JJ;Levine B;Linkermann A;Lipton SA;Lockshin RA;López-Otín C;Lowe SW;Luedde T;Lugli E;MacFarlane M;Madeo F;Malewicz M;Malorni W;Manic G;Marine JC;Martin SJ;Martinou JC;Medema JP;Mehlen P;Meier P;Melino S;Miao EA;Molkentin JD;Moll UM;Muñoz-Pinedo C;Nagata S;Nuñez G;Oberst A;Oren M;Overholtzer M;Pagano M;Panaretakis T;Pasparakis M;Penninger JM;Pereira DM;Pervaiz S;Peter ME;Piacentini M;Pinton P;Prehn JHM;Puthalakath H;Rabinovich GA;Rehm M;Rizzuto R;Rodrigues CMP;Rubinsztein DC;Rudel T;Ryan KM;Sayan E;Scorrano L;Shao F;Shi Y;Silke J;Simon HU;Sistigu A;Stockwell BR;Strasser A;Szabadkai G;Tait SWG;Tang D;Tavernarakis N;Thorburn A;Tsujimoto Y;Turk B;Vanden Berghe T;Vandenabeele P;Vander Heiden MG;Villunger A;Virgin HW;Vousden KH;Vucic D;Wagner EF;Walczak H;Wallach D;Wang Y;Wells JA;Wood W;Yuan J;Zakeri Z;Zhivotovsky B;Zitvogel L;Melino G;Kroemer G
通讯作者：
Kroemer G
影响因子：
14.5
作者：
Mohammad RM;Muqbil I;Lowe L;Yedjou C;Hsu HY;Lin LT;Siegelin MD;Fimognari C;Kumar NB;Dou QP;Yang H;Samadi AK;Russo GL;Spagnuolo C;Ray SK;Chakrabarti M;Morre JD;Coley HM;Honoki K;Fujii H;Georgakilas AG;Amedei A;Niccolai E;Amin A;Ashraf SS;Helferich WG;Yang X;Boosani CS;Guha G;Bhakta D;Ciriolo MR;Aquilano K;Chen S;Mohammed SI;Keith WN;Bilsland A;Halicka D;Nowsheen S;Azmi AS
通讯作者：
Azmi AS
DOI：
10.1016/s0736-5748(02)00048-5
发表时间：
2002-06-01
影响因子：
1.8
作者：
Ye, ZC;Sontheimer, H
通讯作者：
Sontheimer, H