1. Aglycosylated antibody-producing mice for aglycosylated antibody-lectin coupled immunoassay for the quantification of tumor markers (ALIQUAT)
- Author
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Eui-Jeon Woo, Sun Hee Kim, Yong-Sam Kim, Jeong-Heon Ko, Nan-Ee Lee, Dae-Yeul Yu, Myung-Il Kim, Sun Min Lee, and Gi-Sang Seong
- Subjects
0301 basic medicine ,Glycosylation ,Mouse ,Medicine (miscellaneous) ,Enzyme-Linked Immunosorbent Assay ,Mice, Transgenic ,Biochemical assays ,General Biochemistry, Genetics and Molecular Biology ,Immunoglobulin G ,Article ,03 medical and health sciences ,0302 clinical medicine ,Genome editing ,Lectins ,medicine ,Biomarkers, Tumor ,Animals ,Humans ,lcsh:QH301-705.5 ,Immunoassay ,Mice, Inbred BALB C ,biology ,medicine.diagnostic_test ,Chemistry ,Protein Stability ,Lectin ,Antibodies, Monoclonal ,Diagnostic markers ,Fucosyltransferases ,Fragment crystallizable region ,In vitro ,Mice, Inbred C57BL ,030104 developmental biology ,HEK293 Cells ,lcsh:Biology (General) ,Biochemistry ,030220 oncology & carcinogenesis ,Monoclonal ,Genetic engineering ,biology.protein ,alpha-Fetoproteins ,Antibody ,General Agricultural and Biological Sciences - Abstract
Targeting aberrant glycoforms has been validated for in vitro cancer diagnostic development, and several assays are currently in routine clinical use. Because N-glycans in Fc region of antibodies show cross-reactivity with various lectins, high-quality aglycosylated antibodies are exceptionally important for immunoassay platform-based quantitative measurements. Previously, aglycosylated antibody acquisition relied on incomplete, uneconomical and onerous enzymatic and chemical methods. Here, we edited four murine immunoglobulin G genes using adenine base-editing and homology-directed recombination (HDR)-mediated gene editing methods to generate aglycosylated antibody-producing mice. Resulting aglycosylated antibodies showed required analytical performances without compromised protein stability. Thus, this aglycosylated monoclonal antibody-lectin coupled immunoassay for the quantification of tumour markers (ALIQUAT) method can provide a robust, versatile and accessible immunoassay platform to quantify specific glycoforms in precision cancer diagnostics. Moreover, the engineered mice can be used as a host to produce various aglycosylated antibodies in a convenient and robust fashion, thereby expanding in vitro diagnostic development opportunities that utilize glycoforms as a disease-specific biomarkers., Lee et al. describe the generation of aglycosylated antibody-producing mice. These aglycosylated antibodies, lacking glycans prevent unwanted interactions with the lectins, and are used as reagents in a tool they developed called ALIQUAT. This aglycosylated antibody and lectin-based immunoassay diagnostic platform can be used to detect disease specific glycan biomarkers.
- Published
- 2020