Development of on‐chip fully automated immunoassay system “μTASWako i30” to measure the changes in glycosylation profiles of alpha‐fetoprotein in patients with hepatocellular carcinoma

Abstract

Glycosylation profiles significantly change during oncogenesis. Aberrant glycosylation can be used as a cancer biomarker in clinical settings. Different glycoforms can be separately detected using lectin affinity electrophoresis and lectin array-based methods. However, most methodologies and procedures need experienced technique to perform the assays and expertise to interpret the results. To apply glycomarkers for clinical practice, a robust assay system with an easy-to-use workflow is required. Wako's μTASWako i30, a fully automated immunoanalyzer, was developed for in vitro diagnostics based on microfluidic technology. It utilizes the principles of liquid-phase binding assay, where immunoreactions are performed in a liquid phase, and electrokinetic analyte transport assay. Capillary electrophoresis on microfluidic chip has enabled the detection of different glycoform types of alpha-fetoprotein (AFP), a serum biomarker for hepatocellular carcinoma. AFP with altered glycosylation can be separated based on the reactivity to Lens culinaris agglutinin on electrophoresis. The glycoform AFP-L3 was reportedly more specific in hepatocellular carcinoma. This assay system can provide a high sensitivity and rapid results in 9 min. The test results for ratio of AFP-L3 to total AFP using μTASWako i30 are correlated with those of conventional methodology. The μTASWako assay system and the technology can be utilized for glycosylation analysis in the postgenomic era.

Vitronectin (Vn) glycosylation patterned by lectin affinity assays—A potent glycoproteomic tool to discriminate plasma Vn from cancer ascites Vn

“…Thus, the commercial lectin array we used appears suitable for high‐throughput screening and profiling of glycosylation and glycoproteins. Accumulating propositions based on microarray technology and lectin affinity are gaining popularity in glycomics and glycoproteomics . Thanks to their diversity and specificity of glycan recognition, lectins can distinguish between linkages or other fine nuances of oligosaccharide, information not easily accessible using mass spectrometry or chromatography methods .…”

Capillary electrophoresis of glycoproteins

“…As indicated previously, detection of glycoproteins using fluorescence in CZE is very limited. In the reviewed period, this type of detection has been mentioned for the determination of the two glycoforms of AFP (-fetoprotein) -AFP-L1 and AFP-L3-as biomarker for prognostic and therapeutic response evaluation of hepatocellular carcinoma [96]. In this commercialized instrument (µ-TASWako i30 from FUJIFILM Wako Chemicals Europe GmbH), the whole analytical process, including immunorecognition of the AFP in the sample, concentration, separation of the glycoform L1 from the L3, and detection, is carried out in a proprietary microchip.…”

“…Several reviews covering different aspects partially overlapping with this subject have been published in the last five years. For example, Terry Phillips have covered the clinical applications of electrophoresis in capillaries and microchips [207], where some works of glycoforms and glycopeptides as disease markers are mentioned [96]. In the book chapter by Zhang and Hage about applications of CE methods in clinical chemistry a few examples for analysis of glycoprotein isoforms are included [208].…”

“…The technology for the TAS was described in a further paper by the same authors [96]. In this work, the system was employed to probe its usefulness for separating and measuring fucosylated -fetoprotein (AFP-L3) and non-fucosylated forms of AFP as a biomarker of hepatocellular carcinoma (HCC).…”

Recent advances in CE and microchip‐CE in clinical applications: 2014 to mid‐2017

“…The idea of a lab‐on‐a‐chip still continues and the development of a clinically oriented device has been reported by Kurosawa and Watanabe who designed the automated “μTASWako i30” immune‐analyzer. This immune‐analyzer was developed to measure changes in glycosylation profiles during oncogenesis.…”