Selective, sensitive and comprehensive detection of immune complex antigens by immune complexome analysis with papain-digestion and elution

Journal of Immunological Methods
Nozomi AibaraKaname Ohyama

Abstract

Comprehensive identification and profiling of antigens in immune complexes (ICs) in biological fluids, such as serum and cerebrospinal fluid, is useful for developing early diagnostic markers and specific treatments for many diseases. We have developed a method, designated "immune complexome analysis", to comprehensively identify the antigens in ICs. In this method, we first purify ICs from biological fluid by using Protein G- or Protein A-coated beads, then these ICs are subjected to tryptic digestion on the beads and subsequent analysis using nano-liquid chromatography-tandem mass spectrometry (nano-LC-MS/MS). We previously used this method to find specific antigens in circulating ICs (CIC-antigens) in serum for autoimmune diseases, infectious disease and cancers. However, this method detects not only CIC-antigens but also antibodies and proteins bound non-specifically to the beads, which restricts the detection of minor peptides released by the digestion of CIC-antigens whose amounts are generally much less than antibodies and the proteins. To selectively detect CIC-antigens with enhanced sensitivity, in this study we compared three methods (Method A, direct tryptic digestion on the beads; Method B, low-pH elution and trypti...Continue Reading

Citations

Aug 7, 2019·International Journal of Molecular Sciences·Yasuyoshi MiyataHideki Sakai
Jun 9, 2020·Analytical Sciences : the International Journal of the Japan Society for Analytical Chemistry·Nozomi AibaraKaname Ohyama

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