ABSTRACT
Antibody pharmaceuticals have become the most popular immunotherapeutic drugs and are often administered with low serum drug dosages. Hence, the development of a highly sensitive method for the quantitative assay of antibody levels is of great importance to individualized therapy. On the basis of the dual signal amplification by the glycan-initiated site-directed electrochemical grafting of polymer chains (glyGPC), we report herein a novel strategy for the amplified electrochemical detection of antibody pharmaceuticals. The target of interest was affinity captured by a DNA aptamer ligand, and then the glycans of antibody pharmaceuticals were decorated with the alkyl halide initiators (AHIs) via boronate cross-linking, followed by the electrochemical grafting of the ferrocenyl polymer chains from the glycans of antibody pharmaceuticals through the electrochemically controlled atom transfer radical polymerization (eATRP). As the glycans can be decorated with multiple AHIs and the grafted polymer chains are composed of tens to hundreds of electroactive tags, the glyGPC-based strategy permits the dually amplified electrochemical detection of antibody pharmaceuticals. In the presence of trastuzumab (Herceptin) as the target, the glyGPC-based strategy achieved a detection limit of 71.5 pg/mL. Moreover, the developed method is highly selective, and the results of the quantitative assay of trastuzumab levels in human serum are satisfactory. Owing to its uncomplicated operation and cost-effectiveness, the glyGPC-based strategy shows great promise in the amplified electrochemical detection of antibody pharmaceuticals.
Subject(s)
Aptamers, Nucleotide , Electrochemical Techniques , Trastuzumab , Electrochemical Techniques/methods , Humans , Trastuzumab/chemistry , Trastuzumab/blood , Aptamers, Nucleotide/chemistry , Limit of Detection , Polysaccharides/chemistry , Biosensing Techniques/methods , Polymers/chemistryABSTRACT
As a product of nonenzymatic glycation, glycated albumin (GA) is a promising serum marker for the short-term glycemic monitoring in patients with diabetes. On the basis of the boronate crosslinking (BCL)-enabled direct labeling of ferrocene (Fc) tags to the nonenzymatically glycated (NEG) sites, we report herein a novel aptamer-based ratiometric electrochemical (apt-REC) platform for the point-of-care (POC) assay of GA. This apt-REC platform is based on the recognition of GA proteins by the methylene blue (MB)-modified aptamer receptors and the labeling of the Fc tags to the NEG sites via the BCL. Using MB as the reference tag and Fc as the quantification tag, the ratio of the oxidation currents (i.e., IFc/IMB) can serve as the yardstick for the ratiometric assay of GA. Due to the presence of tens of the NEG sites, each GA protein can be labeled with tens of quantification tags, permitting the amplified assay in a simple, time-saving, and low-cost manner. The ratiometric signal exhibited a good linear response over the range from 0.1 to 100 µg/mL, with a detection limit of 45.5 ng/mL. In addition to the superior reproducibility and robustness, this apt-REC platform is highly selective (capable of discriminating GA against human serum albumin (HSA)) and applicable to GA assay in serum samples. Due to its low cost, high reproducibility and robustness, simple operation, and high sensitivity and selectivity, this apt-REC platform holds great promise in the POC assay of GA for diabetes management.