Papas Klearchos K, Pisania Anna, Wu Haiyan, Weir Gordon C, Colton Clark K
Department of Chemical Engineering, Massachusetts Institute of Technology, 25 Ames St., Cambridge, Massachusetts 02139, USA.
Biotechnol Bioeng. 2007 Dec 1;98(5):1071-82. doi: 10.1002/bit.21486.
Improvements in pancreatic islet transplantation for treatment of diabetes are hindered by the absence of meaningful islet quality assessment methods. Oxygen consumption rate (OCR) has previously been used to assess the quality of organs and primary tissue for transplantation. In this study, we describe and characterize a stirred microchamber for measuring OCR with small quantities of islets. The device has a titanium body with a chamber volume of about 200 microL and is magnetically stirred and water jacketed for temperature control. Oxygen partial pressure (pO(2)) is measured by fluorescence quenching with a fiber optic probe, and OCR is determined from the linear decrease of pO(2) with time. We demonstrate that measurements can be made rapidly and with high precision. Measurements with betaTC3 cells and islets show that OCR is directly proportional to the number of viable cells in mixtures of live and dead cells and correlate linearly with membrane integrity measurements made with cells that have been cultured for 24 h under various stressful conditions.
缺乏有意义的胰岛质量评估方法阻碍了用于治疗糖尿病的胰岛移植技术的改进。耗氧率(OCR)此前已被用于评估移植器官和原代组织的质量。在本研究中,我们描述并表征了一种用于测量少量胰岛OCR的搅拌微室。该装置主体由钛制成,腔室体积约为200微升,通过磁力搅拌并带有水套以控制温度。氧分压(pO₂)通过光纤探头的荧光猝灭进行测量,OCR则根据pO₂随时间的线性下降来确定。我们证明测量可以快速且高精度地进行。对βTC3细胞和胰岛的测量表明,OCR与活细胞和死细胞混合物中活细胞的数量成正比,并且与在各种应激条件下培养24小时的细胞的膜完整性测量值呈线性相关。