Quantitative microplate-based respirometry with correction for oxygen diffusion.
Quantitative microplate-based respirometry with correction for oxygen diffusion.
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DOI：
10.1021/ac900881z
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发表时间：
2009-08-15
影响因子：
7.4
通讯作者：
Brand, Martin D.
中科院分区：
文献类型：
--
作者：
Gerencser, Akos A.;Neilson, Andy;Choi, Sung W.;Edman, Ursula;Yadava, Nagendra;Oh, Richard J.;Ferrick, David A.;Nicholls, David G.;Brand, Martin D.
使用改良细胞培养微孔板的呼吸测定法增加了通量，减少了每次测定所需的生物材料。板式呼吸计易受一系列扩散现象的影响;当样本消耗O2时，大气中的O2会泄漏到测量体积中。氧气也溶解在通常用作微孔板材料的聚苯乙烯中并通过聚苯乙烯被动扩散。因此，这种呼吸计腔室的壁不仅对O2是可渗透的，而且还储存大量的气体。壁和测量体积之间的O2通量根据实际[O2]梯度使测得的耗氧率偏置。我们描述了一种基于房室模型的校正算法，从测得的[O2]去卷积的生物耗氧率。我们优化的算法与海马XF 24细胞外通量分析仪。使用小鼠皮质突触体和肝线粒体连接到XF 24 V7细胞培养微板，并通过与经典的克拉克电极血氧仪测量比较，生物学验证校正算法。该算法增加了耗氧率的有用范围、时间分辨率和测量持续时间。该算法以通用格式呈现，因此适用于其他呼吸计系统。
Respirometry using modified cell culture microplates offers an increase in throughput and a decrease in biological material required for each assay. Plate based respirometers are susceptible to a range of diffusion phenomena; as O2 is consumed by the specimen, atmospheric O2 leaks into the measurement volume. Oxygen also dissolves in and diffuses passively through the polystyrene commonly used as a microplate material. Consequently the walls of such respirometer chambers are not just permeable to O2 but also store substantial amounts of gas. O2 flux between the walls and the measurement volume biases the measured oxygen consumption rate depending on the actual [O2] gradient. We describe a compartment model-based correction algorithm to deconvolute the biological oxygen consumption rate from the measured [O2]. We optimize the algorithm to work with the Seahorse XF24 extracellular flux analyzer. The correction algorithm is biologically validated using mouse cortical synaptosomes and liver mitochondria attached to XF24 V7 cell culture microplates, and by comparison to classical Clark electrode oxygraph measurements. The algorithm increases the useful range of oxygen consumption rates, the temporal resolution, and durations of measurements. The algorithm is presented in a general format and is therefore applicable to other respirometer systems.
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影响因子：
3.8
作者：
Dykens, James A.;Jamieson, Joseph D.;Will, Yvonne
通讯作者：
Will, Yvonne
影响因子：
5.5
作者：
Wu, Min;Neilson, Andy;Ferrick, David A.
通讯作者：
Ferrick, David A.
影响因子：
4.7
作者：
BRADFORD, HF
通讯作者：
BRADFORD, HF
影响因子：
4.7
作者：
RAFALOWSKA, U;ERECINSKA, M;WILSON, DF
通讯作者：
WILSON, DF
影响因子：
4.7
作者：
Choi SW;Gerencser AA;Nicholls DG
通讯作者：
Nicholls DG