In the present paper, we describe the fabrication and operation of a FET-based glucose biosensor using glucose oxidase immobilized onto multilayered LbL films of cationic PPI and anionic NiTsPc. Relevant parameters of protocol the films�� preparation and enzyme immobilization were examined and optimized previously [10]. The results showed that the presence of bovine serum albumin (BSA) increases the ENFET signal, and that parameters such as pH and ionic strength are also relevant. The relationship between the output voltage and glucose concentration demonstrates that PPI/NiTsPc-GOx Inhibitors,Modulators,Libraries FET-based biosensor is able to detect glucose.2.?Experimental Section2.1. ChemicalsGlucose oxidase (EC 1.1.3.4 type VII) from Aspergillus niger having 100 units/mg of activity, serum bovine albumin (BSA), NiTsPc and glutaraldehyde (GA) were purchased from Sigma Aldrich and used without purification.
PPI dendrimer (generation 3) was synthesized by a divergent route from an ethylenediamine (EDA) core as described Inhibitors,Modulators,Libraries elsewhere [10]. All other reagents were of analytical grade and used as received. ITO coated glasses (160 nm) were purchased from Delta Technologies and were Inhibitors,Modulators,Libraries cleaned by immersion in a mixture of HNO3-HCl-H2O (1:3:20) for 10 min, followed by washing in Milli-Q water (18.3 M��?cm).2.2. PPI/NiTsPc Growth and GOx ImmobilizationPPI/NiTsPc multilayers were assembled onto ITO substrates by immersing the substrates in polycationic PPI solution (1 mg?mL?1) for 5 min and anionic NiTsPc solution (0.5 mg?mL?1) for 3 min. After each immersion, the pre-coated ITO film was washed with Milli-Q water for 10 seconds and dried under a nitrogen flow.
Five bilayers of PPI/NiTsPc were achieved based on our previous work upon repetition of the cycle described above. Inhibitors,Modulators,Libraries More details of the PPI/NiTsPc growth and characterization can be found elsewhere [10]. GOx was cross-linked on the five PPI/NiTsPc bilayers with a last layer composed of PPI having NH3+ terminated groups by dropping 10 ��L of a mixture of GOx, BSA and GA. For this, 100 ��L of glutaraldehyde (2.5% in water) was mixed with 240 ��L of a mixture containing 20 mg?mL?1 of BSA and 50 mg?mL?1 of GOx according Batimastat to the previously described methodology [11]. Another membrane without BSA was also tested.2.3. FET-Based Biosensor Measurement SystemThe FET-based biosensor device was composed of a biochemically sensitive membrane formed by PPI/NiTsPc-GOx as enzyme separative extended gate, connected to a commercial AD620 amplifier used here as unit gain buffer.
The biomembrane was immersed in a phosphate buffer solution (10 mM, pH 7.5) and glucose aliquots were then added in the measurement cell to determine the glucose sensing characteristics. For the measurements of the time dependence on the output voltage we used a Ag/AgCl/Sat-KCl reference www.selleckchem.com/products/Sorafenib-Tosylate.html electrode to support a constant voltage.