Push-pull perfusion sampling with segmented flow for high temporal and spatial resolution in vivo chemical monitoring.
Push-pull perfusion sampling with segmented flow for high temporal and spatial resolution in vivo chemical monitoring.
复制标题
DOI：
10.1021/ac2003938
复制
发表时间：
2011-07-01
影响因子：
7.4
通讯作者：
Kennedy, Robert T.
中科院分区：
文献类型：
--
作者：
Slaney, Thomas R.;Nie, Jing;Hershey, Neil D.;Thwar, Prasanna K.;Linderman, Jennifer;Burns, Mark A.;Kennedy, Robert T.
低流量推拉灌注是一种采样方法，比竞争性方法（如微透析）产生更好的空间分辨率。由于所使用的低流速（50 nL/min），以高时间分辨率使用该技术是具有挑战性的，这需要收集、操纵和分析纳升样品的方法。高时间分辨率还需要在采样期间控制泰勒色散。为了应对这些挑战，推拉灌注与分段流相结合，以在50 nL/min的流速下以7 s的时间分辨率实现体内采样。通过进一步优化探针入口，在体外证明了以30 nL/min（仅拉动）的200 ms分辨率进行采样。使用这种方法，L-谷氨酸监测在麻醉大鼠的纹状体。以7 s的间隔收集多达500个样品，每个样品6 nL，用不混溶的油分段并储存在毛细管中。通过将样品泵入由Teflon制成的试剂添加三通管中，以15个样品/分钟的速率离线测定样品的L-谷氨酸盐，在该试剂添加三通管中添加试剂用于荧光酶测定。在下游监测所得堵塞物的荧光。微量注射生理缓冲盐水中的70 mM钾诱发L-谷氨酸浓度瞬变，平均最大值为4.5 ± 1.1 μM（n = 6只动物，每只注射3-4次），上升时间为22 ± 2 s。这些结果表明，低流量推挽灌注与分段流可用于高时间分辨率的化学监测和复杂的生物环境。
Low-flow push-pull perfusion is a sampling method that yields better spatial resolution than competitive methods like microdialysis. Because of the low flow rates used (50 nL/min) it is challenging to use this technique at high temporal resolution which requires methods of collecting, manipulating, and analyzing nanoliter samples. High temporal resolution also requires control of Taylor dispersion during sampling. To meet these challenges, push-pull perfusion was coupled with segmented flow to achieve in vivo sampling at 7 s temporal resolution at 50 nL/min flow rates. By further miniaturizing the probe inlet, sampling with 200 ms resolution at 30 nL/min (pull only) was demonstrated in vitro. Using this method, L-glutamate was monitored in the striatum of anesthetized rats. Up to 500 samples of 6 nL each were collected at 7 s intervals, segmented by an immiscible oil and stored in a capillary tube. The samples were assayed offline for L-glutamate at a rate of 15 samples/min by pumping them into a reagent addition tee fabricated from Teflon where reagents were added for a fluorescent enzyme assay. Fluorescence of the resulting plugs was monitored downstream. Microinjection of 70 mM potassium in physiological buffered saline evoked L-glutamate concentration transients that had an average maxima of 4.5 ± 1.1 μM (n = 6 animals, 3–4 injections each) and rise times of 22 ± 2 s. These results demonstrate that low-flow push-pull perfusion with segmented flow can be used for high temporal resolution chemical monitoring and in complex biological environments.
登录
查看更多内容
影响因子：
7.4
作者：
Roman, Gregory T.;Wang, Meng;Shultz, Kristin N.;Jennings, Colin;Kennedy, Robert T.
通讯作者：
Kennedy, Robert T.
影响因子：
7.4
作者：
Kulagina, NV;Shankar, L;Michael, AC
通讯作者：
Michael, AC
影响因子：
4.2
作者：
Patterson, Eric E., II;Pritchett, Jeanita S.;Shippy, Scott A.
通讯作者：
Shippy, Scott A.
DOI：
10.1098/rspa.1956.0065
发表时间：
1956-01-01
影响因子：
--
作者：
ARIS, R
通讯作者：
ARIS, R
影响因子：
7.4
作者：
Huynh, BH;Fogarty, BA;Lunte, SM
通讯作者：
Lunte, SM