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@ARTICLE{CardonaGloria:278419,
      author       = {Y. Cardona Gloria and A. N. R. Weber$^*$},
      title        = {{I}nflammasome {A}ctivation in {H}uman {M}acrophages:
                      {IL}-1β {C}leavage {D}etection by {F}ully {A}utomated
                      {C}apillary-{B}ased {I}mmunoassay.},
      journal      = {Methods in molecular biology},
      volume       = {2696},
      issn         = {1064-3745},
      address      = {[Heidelberg]},
      publisher    = {[Springer]},
      reportid     = {DKFZ-2023-01653},
      isbn         = {978-1-0716-3349-6 (print)},
      pages        = {239-256},
      year         = {2023},
      note         = {Second Edition},
      abstract     = {Interleukin (IL)-1β is a key mediator of inflammation and
                      activates via pattern recognition receptors (PRR) of the
                      inflammasome family by proteolytic maturation. Proteolysis
                      is driven by proteases such as caspase-1 (also known as IL-1
                      converting enzyme, ICE) and converts the intact pro-IL-1β
                      ~31 kDa pro-peptide into a mature, ~17 kDa form that can
                      exit cells through nanomolecular pores or via microvesicles.
                      Whereas pro-IL-1β fails to trigger IL-1 receptor (IL-1R)
                      activation, mature IL-1β, upon release from the cell,
                      triggers pleiotropic downstream effects, establishing an
                      inflammatory state. Hence, being able to detect IL-1β
                      conversion is physiologically relevant for measuring
                      inflammation, but it cannot be easily accomplished by
                      conventional ELISA or flow cytometry as most commercially
                      available antibodies do not discriminate mature and
                      pro-form. Furthermore, unlike for other cytokines, the mere
                      induction and translation of IL1B mRNA cannot serve as a
                      proxy of inflammasome PRR activation. Rather the cleavage of
                      IL-1β needs to be verified. Hence, conventional
                      immunoblotting has emerged as the gold standard for
                      demonstrating inflammasome activation as the difference in
                      molecular weight between pro- and mature form can easily be
                      detected. However, conventional immunoblotting suffers from
                      poor standardization, quantification, and reproducibility,
                      may require sample concentration, and is also not suitable
                      for medium to high throughput. Some of these shortcomings
                      are prohibitive for analysis of human primary samples but
                      can be overcome by fully automated capillary-based
                      immunoassay as we outline here. We here provide a practical
                      guide to quantify pro- vs mature IL-1β directly from
                      unconcentrated supernatants of human monocyte-derived
                      macrophages. The assay may be useful for more standardized
                      and medium-throughput analysis in these cells or other
                      biospecimen.},
      keywords     = {Humans / Inflammasomes / NLR Family, Pyrin
                      Domain-Containing 3 Protein / Reproducibility of Results /
                      Cells, Cultured / Macrophages / Interleukin-1beta /
                      Immunoblotting / Inflammation / Caspase 1 / Automated
                      capillary-based immunoassay (Other) / Caspase-1 (Other) /
                      Human macrophages (Other) / IL-1β (Other) / Inflammasome
                      (Other) / NLRP3 (Other) / Inflammasomes (NLM Chemicals) /
                      NLR Family, Pyrin Domain-Containing 3 Protein (NLM
                      Chemicals) / Interleukin-1beta (NLM Chemicals) / Caspase 1
                      (NLM Chemicals)},
      cin          = {TU01},
      ddc          = {570},
      cid          = {I:(DE-He78)TU01-20160331},
      pnm          = {899 - ohne Topic (POF4-899)},
      pid          = {G:(DE-HGF)POF4-899},
      typ          = {PUB:(DE-HGF)3 / PUB:(DE-HGF)16},
      pubmed       = {pmid:37578727},
      doi          = {DOI:10.1007/978-1-0716-3350-2_16},
      url          = {https://inrepo02.dkfz.de/record/278419},
}