biostudies-literatureTopping JSenior Fellowship from Kennedy TrustCancer Research UKNational Institute for Health Research (NIHR)EU Horizon 2020 research and innovation programmeLeeds Biomedical Research Centreuxae117https://www.ebi.ac.uk/biostudies/studies/S-EPMC11754646biostudies-literatureUnknown219(1)<h4>Introduction</h4>The apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC) is crucial for inflammasome assembly and activation of several inflammasomes, including NLRP3. ASC aggregates are detected in human sera post pyroptotic cell death, but their inflammasome origin remains unclear.<h4>Method</h4>This study aimed to develop a method to detect ASC aggregates originating from NLRP3 inflammasomes. Initially, human monocytes, macrophages, and THP-1 ASC reporter cells were employed to validate the detection of ASC/NLRP3-positive events through flow cytometry.<h4>Results</h4>The presence of ASC/NLRP3 specks was confirmed in cell supernatants from monocytes and macrophages treated with LPS and nigericin or ATP. Flow cytometry analysis identified double-positClinical and experimental immunologyFACS-based detection of extracellular ASC specks from NLRP3 inflammasomes in inflammatory diseases.PMC11754646779295A29685Ibbotson ALara-Reyna SChang LTopping JPeckham DMcDermott MFJarosz-Griffiths HPoulter JSavic SfalseFACS-based detection of extracellular ASC specks from NLRP3 inflammasomes in inflammatory diseases.<h4>Introduction</h4>The apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC) is crucial for inflammasome assembly and activation of several inflammasomes, including NLRP3. ASC aggregates are detected in human sera post pyroptotic cell death, but their inflammasome origin remains unclear.<h4>Method</h4>This study aimed to develop a method to detect ASC aggregates originating from NLRP3 inflammasomes. Initially, human monocytes, macrophages, and THP-1 ASC reporter cells were employed to validate the detection of ASC/NLRP3-positive events through flow cytometry.<h4>Results</h4>The presence of ASC/NLRP3 specks was confirmed in cell supernatants from monocytes and macrophages treated with LPS and nigericin or ATP. Flow cytometry analysis identified double-posit2025-01-01T00:00:00Z2025 Jan2025-04-19T18:20:54.15Z2025-04-19T18:20:54.15ZS-EPMC117546463965768510.1093/cei/uxae117