Mădăraş M B, Spokane R B, Johnson J M, Woodward J R
Department of Chemistry, University of North Carolina at Chapel Hill 27599-3290, USA.
Anal Chem. 1997 Sep 15;69(18):3674-8. doi: 10.1021/ac970173f.
Glutamine membranes for amperometric measurements are described. The interference of the endogenous glutamate is greatly diminished by using a supplementary "anti-glutamate" layer consisting of immobilized glutamate oxidase and catalase on top of the glutamine-sensitive layer having co-immobilized glutaminase and glutamate oxidase. The use of polycarbonate membranes with different permeability characteristics for the control of the substrate's access to the enzyme layers is presented, as well as the effect of the density of the enzyme layer on the sensitivity of these membranes. The fabricated membranes have good operational stability (at least 5 days) and very good linearity (up to 10 mM glutamine). Using an appropriate choice of membranes and cross-linking conditions, membranes with good rejection of glutamate have been fabricated (less than 6% RSD for a 5 mM glutamine sample containing 5 mM glutamate as interferent). These membranes are suitable for monitoring of glutamine levels in mammalian cell cultures without the need of a separate measurement for glutamate.
描述了用于安培测量的谷氨酰胺膜。通过在共固定谷氨酰胺酶和谷氨酸氧化酶的谷氨酰胺敏感层顶部使用由固定化谷氨酸氧化酶和过氧化氢酶组成的补充“抗谷氨酸”层，可大大减少内源性谷氨酸的干扰。介绍了使用具有不同渗透特性的聚碳酸酯膜来控制底物进入酶层，以及酶层密度对这些膜灵敏度的影响。所制备的膜具有良好的操作稳定性（至少5天）和非常好的线性（高达10 mM谷氨酰胺）。通过适当选择膜和交联条件，制备出了对谷氨酸具有良好排斥性的膜（对于含有5 mM谷氨酸作为干扰物的5 mM谷氨酰胺样品，相对标准偏差小于6%）。这些膜适用于监测哺乳动物细胞培养物中的谷氨酰胺水平，而无需单独测量谷氨酸。
