Journal of immunological methods · 2026
Development of chimeric DGP-IgG antibodies as quality control for celiac disease diagnosis.
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Zheng Liang, Lin Xiaotao, Xiang Xufu, Qian Chungen, Cheng Fangming, Wang Gang, He Yuxi, Tan Hui
Аннотация
BACKGROUND: Celiac disease (CD) is an autoimmune disorder triggered by an immune response to dietary gluten in the small intestine, which is frequently undiagnosed or misdiagnosed owing to its non-specific symptoms. The detection of specific serum antibodies, including anti-tissue transglutaminase (tTG) IgA and anti-deamidated gliadin peptide (DGP) IgG, is essential for diagnosing celiac disease. However, the limited availability of DGP IgG positive human sera for use as quality control materials, particularly in countries with low celiac disease prevalence, poses a significant challenge.
OBJECTIVE: To overcome the limited supply of positive serum for DGP IgG, chimeric human DGP IgG antibodies were developed as alternative quality control materials (first time in China). Additionally, several in-house DGP IgG-specific assays (ELISA and CLIA) targeting human and mouse antibodies were developed to monitor each step during chimeric antibodies preparation as quality control materials.
METHODS: Mouse monoclonal antibodies against DGP IgG were screened from hybridoma cells derived from mice immunized with recombinant DGP. Chimeric antibodies of DGP IgG comprising murine variable regions and human IgG constant regions were produced in mammalian expression system using HEK293F cells. Four in-house DGP IgG specific assays were developed to monitor each step of chimeric antibody preparation, i.e. DGP IgG ELISA (mouse), DGP IgG CLIA (mouse), DGP IgG ELISA (human) and DGP IgG CLIA (human). Final chimeric antibodies were prepared as quality control materials and tested for homogeneity and open-vial stability. SPR (Surface Plasmon Resonance) was used to measure binding affinity for DGP chimeric antibodies and DGP antigen for DGP IgG assays mentioned above.
RESULTS: The chimeric antibodies against DGP demonstrated high affinity to DGP antigen by using SPR analysis. The chimeric DGP IgG as quality materials exhibited satisfactory homogeneity (CV < 10%) and open-vial stability (for at least 5 days at 2-8 °C and at least 90 days at -20 °C).
CONCLUSION: This study represents the first development of chimeric DGP IgG as quality control materials and four in-house DGP IgG assays in China. These DGP IgG quality control materials and in-house DGP IgG assays can serve as valuable tools for future DGP IgG assay development and standardization.
OBJECTIVE: To overcome the limited supply of positive serum for DGP IgG, chimeric human DGP IgG antibodies were developed as alternative quality control materials (first time in China). Additionally, several in-house DGP IgG-specific assays (ELISA and CLIA) targeting human and mouse antibodies were developed to monitor each step during chimeric antibodies preparation as quality control materials.
METHODS: Mouse monoclonal antibodies against DGP IgG were screened from hybridoma cells derived from mice immunized with recombinant DGP. Chimeric antibodies of DGP IgG comprising murine variable regions and human IgG constant regions were produced in mammalian expression system using HEK293F cells. Four in-house DGP IgG specific assays were developed to monitor each step of chimeric antibody preparation, i.e. DGP IgG ELISA (mouse), DGP IgG CLIA (mouse), DGP IgG ELISA (human) and DGP IgG CLIA (human). Final chimeric antibodies were prepared as quality control materials and tested for homogeneity and open-vial stability. SPR (Surface Plasmon Resonance) was used to measure binding affinity for DGP chimeric antibodies and DGP antigen for DGP IgG assays mentioned above.
RESULTS: The chimeric antibodies against DGP demonstrated high affinity to DGP antigen by using SPR analysis. The chimeric DGP IgG as quality materials exhibited satisfactory homogeneity (CV < 10%) and open-vial stability (for at least 5 days at 2-8 °C and at least 90 days at -20 °C).
CONCLUSION: This study represents the first development of chimeric DGP IgG as quality control materials and four in-house DGP IgG assays in China. These DGP IgG quality control materials and in-house DGP IgG assays can serve as valuable tools for future DGP IgG assay development and standardization.