A Pencil-Lead Immunosensor for the Rapid Electrochemical Measurement of Anti-Diphtheria Toxin Antibodies

Abstract

Diphtheria is a vaccine-preventable disease, yet immunization can wane over time to non-protective levels. We have developed a low-cost, miniaturized electroanalytical biosensor to quantify anti-diphtheria toxin (DTx) immunoglobulin G (anti-DTx IgG) antibody to minimize the risk for localized outbreaks. Two epitopes specific to DTx and recognized by antibodies generated post-vaccination were selected to create a bi-epitope peptide, biEP, by synthesizing the epitopes in tandem. The biEP peptide was conjugated to the surface of a pencil-lead electrode (PLE) integrated into a portable electrode holder. Captured anti-DTx IgG was measured by square wave voltammetry from the generation of hydroquinone (HQ) from the resulting immunocomplex. The performance of the biEP reagent presented high selectivity and specificity for DTx. Under the optimized working conditions, a logarithmic calibration curve showed good linearity over the concentration range of 10−5–10−1 IU mL−1 and achieved a limit of detection of 5 × 10−6 IU mL−1. The final device proved suitable for interrogating the immunity level against DTx in actual serum samples. Results showed good agreement with those obtained from a commercial enzyme-linked immunosorbent assay. In addition, the flexibility for conjugating other capture molecules to PLEs suggests that this technology could be easily adapted to the diagnoses of other pathogens.

Rapid Detection of Anti-SARS-CoV-2 Antibodies With a Screen-Printed Electrode Modified With a Spike Glycoprotein Epitope

“…First, the one-way ANOVA test was performed with a significance of 95% to check whether the device-to-device surface variation during the optimization of preparation or analytical parameters -e.g., sequence of EP, sec-IgG and EP concentrations, and serum dilution ratio -was significant, as done in studies found in the literature [7,17,18]. Next, a two-tailed Students t-test with a confidence level of 95% was performed for pairwise comparisons [7,17,18].…”

“…Serological diagnostic tests that utilize the presence of antibody as the definition for if an individual was infected by a pathogen detects antibodies in their serum or blood. This can be conducted as a lab-based assay such as an enzyme-linked immunosorbent assay (ELISA) [7] or chemiluminescent immunoassay [22], which are time-consuming and expensive, or a point-of-care test based on lateral flow technology that can show limited sensitivity [23,24]. We propose to implement an electrochemical assay based on commercially available SPEs that can be easily sensitized to capture diagnostic antibodies.…”

“…Generally, an electrochemical immunosensor consists of an electron-conducting solid surface where the molecule of interest (antibody or antigen) can be selectively captured, its presence detectd by tthe amplification [7] or suppression [25] of an electrochemical signal. A key element of the biosensor is the biological component that can be immobilized onto its surface such as antigen [25] or EP [7]. Here, the SPE was modified with a peptide representing an EP specific to the S protein of SARS-CoV-2.…”

“…As a platform, electrochemical biosensors stand out to meet the demands for serological diagnostics through their characteristics; rapid, simple, portable, sensitive, easy-to-use, miniaturize, and compatibility with portable instruments [6][7][8][9]. They can be combined with a wide range of biological recognition elements to detect clinically relevant compounds such as enzymes, nucleic acid, antibodies, epitopes, and others [10][11][12][13][14][15].…”

“…They can be combined with a wide range of biological recognition elements to detect clinically relevant compounds such as enzymes, nucleic acid, antibodies, epitopes, and others [10][11][12][13][14][15]. Amongst these choices, epitopes (Eps) hold a high interest since they represent the minimum amino acid sequences in a pathogens proteome that are bound by antibodies generated in a patient in response to an infection [7]. Their use can improve selectivity by the elimination of cross-reactivity based on sequence similarity to other pathogen, which a major concern when whole antigens are used due to the presence of non-specific epitopes that can react with antibodies [16].…”