Amplification-Free Ratiometric Electrochemical Aptasensor for Point-of-Care Detection of Therapeutic Monoclonal Antibodies

Abstract

The rapid quantification of therapeutic monoclonal antibodies (mAbs) is of great significance to their pharmacokinetics/pharmacodynamics (PK/PD) research and the personalized medication for disease treatment. Taking advantage of the direct decoration of tens of redox tags to the target of interest, we illustrate herein an amplification-free ratiometric electrochemical aptasensor for the point-of-care (POC) detection of trace amounts of therapeutic mAbs. The POC detection of therapeutic mAbs involved the use of the methylene blue (MB)-conjugated aptamer as the affinity element and the decoration of therapeutic mAbs with ferrocene (Fc) tags via the boronate crosslinking, in which the MB-derived peak current was used as the reference signal, and the peak current of the Fc tag was used as the output signal. As each therapeutic mAb carries tens of diol sites for the site-specific decoration of the Fc output tags, the boronate crosslinking enabled the amplification-free detection, which is cost-effective and quite simple in operation. In the presence of bevacizumab (BevMab) as the target, the resulting ratiometric signal (i.e., the I Fc/I MB value) exhibited a good linear response over the range of 0.025–2.5 μg/mL, and the limit of detection (LOD) of the electrochemical aptasensor was 6.5 ng/mL. Results indicated that the aptamer-based affinity recognition endowed the detection of therapeutic mAbs with high selectivity, while the ratiometric readout exhibited satisfactory reproducibility and robustness. Moreover, the ratiometric electrochemical aptasensor is applicable to the detection of therapeutic mAbs in serum samples. Taking together, the amplification-free ratiometric electrochemical aptasensor holds great promise in the POC detection of therapeutic mAbs.

Site-Directed Electrochemical Grafting for Amplified Detection of Antibody Pharmaceuticals

“…After the glyGPC for 30 min at room temperature, the obtained electrode was moderately electrolyzed for 20 s at 0.19 V (vs. saturated calomel electrode (SCE)) to remove the Cu(0) residues. Following a stringent wash with ethanol and Milli-Q water, the square wave voltammetry (SWV) response signals were collected in 25 mM LiClO 4 …”

“…The high specificity and affinity of antibodies have flourished their widespread use in the diagnosis and therapy of human diseases . As for therapeutic use, antibody pharmaceuticals, especially the therapeutic monoclonal antibodies, have emerged as the best-selling biopharmaceutical products in the targeted therapy of human diseases, such as cancers, neurological disorders, infectious diseases, and cardiovascular diseases, owing to their high immune effector functions, exquisite specificity, and long serum half-life. , For example, infliximab and adalimumab targeting tumor necrosis factor alpha (TNF-α) have been approved for the immunotherapy of Crohn’s disease, rheumatoid arthritis, and ulcerative colitis; lecanemab against the amyloid-beta (Aβ) soluble protofibrils is undergoing clinical trials to treat early Alzheimer’s disease; trastuzumab against human epidermal growth factor receptor 2 (HER2), cetuximab against epidermal growth factor receptor (EGFR), and atezolizumab against programmed death-ligand 1 (PD-L1) have been approved for the therapy of breast cancer, colorectal cancer, and urothelial carcinoma, respectively. Nevertheless, the economic burden of antibody therapeutics is very high (e.g., the average cost per month for the treatment of chronic lymphocytic leukemia with ofatumumab is up to $16,000), and the effective serum drug dosages of antibody pharmaceuticals are highly individual-dependent.…”

“…Over the past decades, many methods have been reported for the selective detection of antibody levels. − In addition to liquid chromatography–mass spectrometry (LC–MS)-based methods that do not require the use of an affinity ligand, a variety of methods utilizing antibodies, peptide mimotopes, and aptamers, , as the affinity ligands have been reported, such as the surface plasmon resonance (SPR), chemiluminescent, photoelectrochemical (PEC), bioluminescent, fluorescent, , quartz crystal microbalance (QCM), and electrochemical methods. − Among these methods, the electrochemical aptamer-based (E-AB) detection of antibody pharmaceuticals has received increasing attention, largely due to the good portability, low sample consumption and detection cost, and uncomplicated instrumentation. , On the basis of the binding-induced conformational change of the ferrocene (Fc)-modified DNA aptamer, for example, Nagata et al reported an E-AB method for the one-step detection of bevacizumab, with a detection limit of 9.1 nM . This method is easy to operate, low-cost, and highly selective; however, the “signal-off” detection can result in false-positive responses while the detection sensitivity remains to be further improved.…”

Maintenance-free antifouling polymeric membrane potentiometric sensors based on self-polishing coatings

“…Nowadays, potentiometric polymeric membrane ion-selective electrodes (ISEs) are playing an indispensable role in monitoring of environmental elements such as alkali and alkaline earth-metal cations, ammonium ions, nutrients and chemical species relevant for the carbon cycle as well as heavy metals and organic pollutants in environmental waters owing to their excellent selectivity, rapid response, simplicity, low cost, and high reliability. 1–13 However, when these electrodes are applied in marine environments, especially in long-term monitoring, marine fouling often occurs on the surfaces of the sensors owing to the relatively high hydrophobicity of polymeric electrode membrane matrices. Such fouling on the sensor surface usually involves the following steps: surface adsorption of natural organic macromolecules (proteins, polysaccharides, glycoproteins) from seawater to form a conditioning film, adhesion of bacteria via a cellular appendix and exopolymers to finally form a biofilm, anchoring of multicellular species ( e.g.…”