Development of an Immunosensor for PfHRP 2 as a Biomarker for Malaria Detection

Abstract

Plasmodium falciparum histidine-rich protein 2 (PfHRP 2) was selected in this work as the biomarker for the detection and diagnosis of malaria. An enzyme-linked immunosorbent assay (ELISA) was first developed to evaluate the immunoreagent’s suitability for the sensor’s development. A gold-based sensor with an integrated counter and an Ag/AgCl reference electrode was first selected and characterised and then used to develop the immunosensor for PfHRP 2, which enables a low cost, easy to use, and sensitive biosensor for malaria diagnosis. The sensor was applied to immobilise the anti-PfHRP 2 monoclonal antibody as the capture receptor. A sandwich ELISA assay format was constructed using horseradish peroxidase (HRP) as the enzyme label, and the electrochemical signal was generated using a 3, 3′, 5, 5′tetramethyl-benzidine dihydrochloride (TMB)/H2O2 system. The performance of the assay and the sensor were optimised and characterised, achieving a PfHRP 2 limit of detection (LOD) of 2.14 ng·mL−1 in buffer samples and 2.95 ng∙mL−1 in 100% spiked serum samples. The assay signal was then amplified using gold nanoparticles conjugated detection antibody-enzyme and a detection limit of 36 pg∙mL−1 was achieved in buffer samples and 40 pg∙mL−1 in serum samples. This sensor format is ideal for malaria detection and on-site analysis as a point-of-care device (POC) in resource-limited settings where the implementation of malaria diagnostics is essential in control and elimination efforts.

Development of Electrochemical Nanosensor for the Detection of Malaria Parasite in Clinical Samples

“…The LoDs recorded in this work are higher than 2.5 x 10 −5 μg/mL and 4.0 x 10 −5 μg/mL reported for Plasmodium falciparum histidine-rich protein 2 (PfHRP 2) detection on screen printed gold electrode (Gikunoo et al, 2014; Aver et al, 2017). Although the LoD is higher for the present study, the use of chemical biomarkers such as β-hematin should be encouraged because they are stable, affordable (cheaper), and easily available than PfHRP 2.…”

Recent Advances in the Development of Biosensors for Malaria Diagnosis

“…More recently, Hemben et al used anti-PfHRP-2 monoclonal antibodies to capture PfHRP-2 at the surface of a screen printed gold electrode (Table 2) [60]. The captured antigen was targeted with HRP-labelled antibodies and the quantification of PfHRP-2 derived from the substrate (TMB-H 2 O 2 )-enzyme reaction by amperometry ( Figure 2A).…”

“…Strategy for (A) labelled amperometric[60] and (B) label free impedimetric[57] electrochemical detection of PfHRP-2. (A) (i) Gold nanoparticle amplified sandwich assay and (ii) plot of chronoamperometric response of PfHRP-2 detection in spiked serum (0.05-0.5 ng/mL PfHRP).…”

“…(A) (i) Gold nanoparticle amplified sandwich assay and (ii) plot of chronoamperometric response of PfHRP-2 detection in spiked serum (0.05-0.5 ng/mL PfHRP). Reprinted from Hemben et al[60] with permission from MDPI. (B) (i) CZnONF dispersions dropcasted onto GCE followed by chemisoption of anti-HRP2 unto MPA treated electrodes and (ii) calibration curve of impedimetric responses obtained after incubating GCE/fCuZnONFs/Anti-HRP2 biosensor with varying concentrations of PfHRP2 (10 ag/mL-10 µg/mL).…”

Nanoparticle-based Point of Care Immunoassays for in vitro Biomedical Diagnostics

“…2(d)). 45 AuNP-based disposable DNA biosensor strips have also been reported for quick and easy detection of food borne pathogens such as Vibrio cholerae. AuNPs (40 nm) coupled with antifluorescein antibodies were employed to form a detector reagent in a glass fiber-based DNA biosensor for the detection of biotin and fluorescein labeled PCR amplified target.…”