Nanobiotechnology advanced antifouling surfaces for the continuous electrochemical monitoring of glucose in whole blood using a lab-on-a-chip

Abstract

In the current work we have developed a lab-on-a-chip containing embedded amperometric sensors in four microreactors that can be addressed individually and that are coated with crystalline surface protein monolayers to provide a continuous, stable, reliable and accurate detection of blood glucose. It is envisioned that the microfluidic device will be used in a feedback loop mechanism to assess natural variations in blood glucose levels during hemodialysis to allow the individual adjustment of glucose. Reliable and accurate detection of blood glucose is accomplished by simultaneously performing (a) blood glucose measurements, (b) autocalibration routines, (c) mediator-interferences detection, and (d) background subtractions. The electrochemical detection of blood glucose variations in the absence of electrode fouling events is performed by integrating crystalline surface layer proteins (S-layer) that function as an efficient antifouling coating, a highly-oriented immobilization matrix for biomolecules and an effective molecular sieve with pore sizes of 4 to 5 nm. We demonstrate that the S-layer protein SbpA (from Lysinibacillus sphaericus CCM 2177) readily forms monomolecular lattice structures at the various microchip surfaces (e.g. glass, PDMS, platinum and gold) within 60 min, eliminating unspecific adsorption events in the presence of human serum albumin, human plasma and freshly-drawn blood samples. The highly isoporous SbpA-coating allows undisturbed diffusion of the mediator between the electrode surface, thus enabling bioelectrochemical measurements of glucose concentrations between 500 μM to 50 mM (calibration slope δI/δc of 8.7 nA mM(-1)). Final proof-of-concept implementing the four microfluidic microreactor design is demonstrated using freshly drawn blood. Accurate and drift-free assessment of blood glucose concentrations (6. 4 mM) is accomplished over 130 min at 37 °C using immobilized enzyme glucose oxidase by calculating the difference between autocalibration (10 mM glc) and background measurements. The novel combination of biologically-derived nanostructured surfaces with microchip technology constitutes a powerful new tool for multiplexed analysis of complex samples.

Microfluidic enzymatic biosensing systems: A review

“…Different from oxygen, they can be co-immobilized with the enzyme or added to the sample solution in excess so that shortages are avoided and the linear range extended. Picher et al examined the redox mediators hydroquinone, ferricyanide, 8-hydroxyquinoline, anthraquinone and menadione for their compatibility with glucose oxidase and signal strength of the amperometric read out circuit (Picher et al, 2013). Ferricyanide was found to have the best biocompatibility and give the strongest measurement signal.…”

Automated, Miniaturized, and Integrated Quality Control-on-Chip (QC-on-a-Chip) for Cell-Based Cancer Therapy Applications

“…Routinely used immunofluorescence end-point analysis can now be extended with complementary, continuous monitoring techniques (Charwat et al, 2013a(Charwat et al, ,b, 2014Picher et al, 2013;Novak et al, 2015). Several portable and miniaturized sensors have been developed over the past years to analyze rapidly changing biological systems.…”

Biomimetic interfaces based on S-layer proteins, lipid membranes and functional biomolecules

“…One important feature of S-layer proteins is the capability of isolated native or recombinantly produced subunits to selfassemble into crystalline arrays on many surfaces such as glass, silicon oxide and nitride, mica, noble metals like gold, titanium and platinum, but also on stainless steel or many polymers such as polystyrene, polyester and cellulose, and on technically relevant surfaces like highly oriented pyrolytic graphite or indium tin oxide [92,93]. Transmission electron microscopy (TEM) [94][95][96] and atomic force microscopy (AFM) [97][98][99][100][101] are the most appropriate techniques to elucidate the recrystallization process of S-layer proteins.…”