A novel mouse monoclonal antibody c42 against c-terminal peptide of alpha-1-antitrypsin

Srinu Tumpara, Elena Korenbaum, Mark Kühnel, Danny Jonigk, Beata Olejnicka, Michael Davids, Tobias Welte, Beatriz Martinez-Delgado, Sabina Janciauskiene

Research output: Contribution to journalArticlepeer-review

Abstract

The C-terminal-fragments of alpha1-antitrypsin (AAT) have been identified and their diverse biological roles have been reported in vitro and in vivo. These findings prompted us to develop a monoclonal antibody that specifically recognizes C-36 peptide (corresponding to residues 359–394) resulting from the protease-associated cleavage of AAT. The C-36-targeting mouse mono-clonal Immunoglobulin M (IgM) antibody (containing κ light chains, clone C42) was generated and enzyme-linked immunosorbent assay (ELISA)-tested by Davids Biotechnologie GmbH, Germany. Here, we addressed the effectiveness of the novel C42 antibody in different immunoassay formats, such as dot-and Western blotting, confocal laser microscopy, and flow cytometry. According to the dot-blot results, our novel C42 antibody detects the C-36 peptide at a range of 0.1–0.05 µg and shows no cross-reactivity with native, polymerized, or oxidized forms of full-length AAT, the AAT-elastase complex mixture, as well as with shorter C-terminal fragments of AAT. However, the C42 antibody does not detect denatured peptide in SDS-PAGE/Western blotting assays. On the other hand, our C42 antibody, unconjugated as well as conjugated to DyLight488 fluorophore, when applied for immunofluorescence microscopy and flow cytometry assays, specifically detected the C-36 peptide in human blood cells. Altogether, we demonstrate that our novel C42 antibody successfully recognizes the C-36 peptide of AAT in a number of immunoassays and has potential to become an important tool in AAT-related studies.

Original languageEnglish
Article number2141
Number of pages16
JournalInternational Journal of Molecular Sciences
Volume22
Issue number4
DOIs
Publication statusPublished - 2021

Subject classification (UKÄ)

  • Cell and Molecular Biology

Keywords

  • Alpha1-antitrypsin
  • C-terminal peptide
  • C42 clone
  • Immunoassays
  • Molecular forms
  • Monoclonal antibody

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