The development of electrochemical biosensors for cholesterol

Ahmadraji, Termeh

Abstract

Coronary vascular disease (CVD) is the number one cause of death worldwide. According to a WHO report and global and regional projections of mortality and burden of disease, by 2030, the number of people dying from heart disease and stroke will increase to reach 23.3 million. Non-HDL cholesterol, determined by subtracting the high density lipoprotein cholesterol (HDL-C) concentration from the total cholesterol (TC) content, has been recommended as a target for preliminary CVD prevention. In recent times, single-step homogeneous assays have been developed which allow simple and selective measurement of cholesterol fractions. Electrochemical sensors have also been developed which are based on the electrocatalysis of hydrogen peroxide using low cost printed sensor methodologies and such platforms would have the potential to be used as the basis of fabricating cholesterol biosensors for point of care use.
Here, the development of electrochemical biosensors for the selective measurement of HDL-C, TC, and by subtraction non-HDL-C was explored. A spectrophotometric assay for use at room temperature and with minimal sample dilution was first established in order to optimise the assay reagent components for the development of selective cholesterol assays. Assay chemistries based on polyoxyethylene tribenzylphenyl ethers (Emulgen B-66) and Triton X-100 for the selective measurement of HDL-C and TC, respectively, were developed. The impact of these reagents on the electrocatalytic reduction of hydrogen peroxide at silver paste screen printed electrodes was also evaluated and optimised.
Electrochemical biosensors for HDL-C and TC using externally added assay reagents were developed by combining the homogeneous assay methodologies with the printed electrocatalytic electrodes. The effects of assay reagents such as surfactants, enzymes, HDL-C sample and delipidated serum on the electrode behaviour were assessed amperometrically in the presence of hydrogen peroxide solutions. The electrodes showed increases in their catalytic activity toward hydrogen peroxide in the presence of both selective and non-selective surfactant and decreases in the presence of cholesterol oxidase and HDL-C samples. Despite the negative effects of cholesterol oxidase and sample matrix on electrode behaviour, the electrode response was linear within the clinically relevant ranges of HDL-C and TC. The modified electrodes were evaluated for their ability to selectively measure HDL-C and TC in clinical serum samples. The resulting HDL-C biosensor yielded a sensitivity of 3.32×10-8 A/mM with a linear range of 0 to 4 mM (r2=0.999), LOD of 0.5 mM and average RSD of 9.5% (n=5) while the TC biosensor had a sensitivity of 2.24×10-8 A/mM and a linear range of 0 to 10 mM r2=0.984), LOD of 2 mM and average RSD of 10.8% (n=3). The correlation between the HDL-C sensor and a commercial laboratory assay in clinical serum samples had a slope of 0.87 and a Pearson correlation coefficient of 0.76 (n=13) while the correlation for TC measurement had a slope of 1.07 and a Pearson correlation coefficient of 0.87.
Finally, in order to develop a disposable biosensor suitable for point of care testing, integrated biosensors for HDL-C and TC were fabricated by inkjet-print deposition of assay reagents on the electrode surface. Integrated biosensors for the measurement of HDL-C were optimised and yielded a sensitivity of 4.55×10-8 A/mM with a linear range of 0 to 4 mM (r2=0.993) with an LOD of 0.25 mM and average RSD of 6.6% (n=3). The integrated TC biosensor had a sensitivity of 9.38×10-9 A/mM and linear range of 0 to 9 mM (r2=0.982), LOD of 0.5 mM and average RSD of 9.5% (n=3).

• Thesis Type: Thesis
• Publicly Available Date: Jun 6, 2019
• Keywords: electrochemical biosensors, cholesterol
• Public URL: https://uwe-repository.worktribe.com/output/839336
• Award Date: Jan 21, 2015