Human haptoglobin phenotypes and concentration determination by nanogold-enhanced electrochemical impedance spectroscopy

Abstract

Haptoglobin (Hp) is an acute phase protein that binds free hemoglobin (Hb), preventing Hb-induced oxidative damage in the vascular system. There are three phenotypes in human Hp, whose heterogeneous polymorphic structures and varying concentrations in plasma have been attributed to the cause of diseases and outcome of clinical treatments. Different phenotypes of Hp may be composed of the same subunits but different copy numbers, rendering their determination difficult by a single procedure. In this study, we have developed a simple, fast, reliable and sensitive method, using label-free nanogold-modified bioprobes coupled with self-development electrochemical impedance spectroscopy (EIS). By this method, probe surface charge transfer resistance is detected. The relative charge transfer resistance ratios for Hp 1-1, Hp 2-1 and Hp 2-2 were characterized. We were able to determine protein size difference within 3 nm, and the linear region of the calibration curve for Hp levels in the range of 90 pg ml(-1) and 90 µg ml(-1) (∼1 fM to 1 pM). We surmise that similar approaches can be used to investigate protein polymorphism and altered protein-protein interaction associated with diseases.

Electrochemical sensor for nitric oxide using layered films composed of a polycationic dendrimer and nickel(II) phthalocyaninetetrasulfonate deposited on a carbon fiber electrode

“…This concept is not entirely new. Cheng et al demonstrated that protein polymorphisms can be identified by the charge transfer resistance ratio of the protein-bound electrode from EIS measurement [29]. The authors proposed that a simple detection method based on measurement of relative charge transfer resistance ratio by EIS was a potential approach for investigating polymorphism due to changes in molecular weight [29].…”

Rapid and highly sensitive detection of Enterovirus 71 by using nanogold-enhanced electrochemical impedance spectroscopy

“…The value of R ct , which presents the charge transfer from the electrode during the Faradaic reaction, is the optimal parameter of the elements to analyse the Ag-Ab reaction of the working electrode in the Randles circuit. The nonhomogeneous surface of each probe would limit the application of EIS for routine clinical detection [21]. However, as indicated in the previous description, the relative charge transfer resistance ratio ( R ct(Analyte) /R ct(Ab w/BSA) ) can solve this problem and has been applied in previous studies [22].…”

“…Our previous study demonstrated that the haptoglobin (Hp) detection probe can distinguish phenotypes within 1-3 nm of each other and can quantify accurately down to picomolar levels of Hp 2-2, where the molecular size and weight are 13 nm and 190 kDa respectively [21]. EV71 is an icosahedral structure consisting of 60 copies of four capsid proteins.…”

“…There are many type of SAMs applied as the interface-layer in biosensors, such as CGC, 3-MPA, and 3-MPS [21,27,28]. The detection range may be affected by the conducting feature of the SAM layer.…”

“…When the electrode is modified with AuNPs and a specific Ab, EIS can be an excellent tool to detect trace microorganisms. This powerful technique monitors events on sensing electrodes, and has shown promising results for point-of-care and real-time monitoring [19][20][21].…”