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Journal: Scientific Reports

Article Title: KN3014, a piperidine-containing small compound, inhibits auto-secretion of IL-1β from PBMCs in a patient with Muckle–Wells syndrome

doi: 10.1038/s41598-020-70513-0

Figure Lengend Snippet: High-throughput screening of a 9,600 core chemical library using NLRP3 inflammasome in a cell-free system. ( a ) Schematic representations of C-terminal biotinylated full-length NLRP3 (NLRP3-FL-Btn), the N-terminal FLAG-tagged PYD of ASC (FLAG-ASC-PYD), and the N-terminal FLAG-tagged caspase recruit domain (CARD) of ASC (FLAG-ASC-CARD). PYD pyrin domain, CARD caspase recruitment domain, NOD nucleotide-binding oligomerization domain, LRR leucine rich repeat. ( b ) Schematic representation of reconstituted inflammasomes. The PYD of truncated ASC was able to open and bind to the PYD of NLRP3. The chemical energy of the reactive oxygen on donor beads was transferred to acceptor beads, and a signal was detected. ( c ) Primary screening of the 9,600 core chemical library using NLRP3 inflammasome in a cell-free system. The result presented was the only result obtained.

Article Snippet: In addition, 10 µL of an ASC-PYD mixture containing 1 µL FLAG-ASC-PYD was added to sample wells and positive control wells using a FlexDrop dispenser, and 10 µL of an ASC-CARD mixture containing 1 µL FLAG-ASC-CARD was added to negative control wells using a Picus NxT dispenser (Sartorius, Goettingen, Germany).

Techniques: High Throughput Screening Assay, Binding Assay