Development of an enrichment-free one-pot sample preparation and ultra-high performance liquid chromatography-tandem mass spectrometry method to identify Immunoglobulin A1 hinge region O-glycoforms for Immunoglobulin A nephropathy

Hsiao Fan Chen, Chih Chin Kao, Shuk Man Ka, San Yuan Wang, Michael X. Chen, Guan Yuan Chen, Te I. Weng, Ruo Yun Lai, Shu Ching Yeh, Yen Chung Lin, Hsi Hsien Chen, Wei Chiao Chang, Mai Szu Wu, I. Lin Tsai

Research output: Contribution to journalArticlepeer-review

2 Citations (Scopus)

Abstract

Immunoglobulin A nephropathy (IgAN) is a highly prevalent autoimmune renal disease. Human IgA1 with galactose deficiency in the hinge region (HR) has been identified as an autoantigen for this disease. Therefore, analyzing IgA1 HR glycoforms in biofluids is important for biomarker discovery. Herein, an analytical method that includes one-pot sample preparation with unbiased plasma IgA purification, dual internal standard addition, and sensitive ultra-high-performance liquid chromatography-triple quadrupole tandem mass spectroscopy (UHPLC-QqQ-MS/MS) was developed. Targeted O-glycopeptides detection was performed in pooled plasma with the validation of theoretical retention times, enzymatic treatment outcomes, product ion scans, and signal repeatability. A total of 42 IgA1 O-glycopeptides with N-acetylgalactosamines, galactoses, and sialic acids were determined from 8 µL of plasma. The newly developed method was applied to plasma samples from 16 non-IgAN controls and 19 IgAN patients. Comparing the 42 targets, 16 IgA1 HR O-glycopeptides were statistically different between the two groups (p<0.05). Decreased sialylation was identified in the IgA1 hinge region of IgAN patients, which was also correlated with the estimated glomerular filtration rate (eGFR). The developed method is sensitive and precise and can be used to identify plasma biomarkers for IgA nephropathy.

Original languageEnglish
Article number463589
JournalJournal of Chromatography A
Volume1685
DOIs
Publication statusPublished - Dec 6 2022

Keywords

  • Biomarkers
  • IgA nephropathy
  • Immunoglobulin A
  • Liquid chromatography-mass spectrometry
  • O-glycosylation

ASJC Scopus subject areas

  • Analytical Chemistry
  • Biochemistry
  • Organic Chemistry

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