PMID- 28268194
OWN - NLM
STAT- MEDLINE
DCOM- 20170915
LR  - 20180321
IS  - 1872-7905 (Electronic)
IS  - 0022-1759 (Linking)
VI  - 447
DP  - 2017 Aug
TI  - A bioluminescent caspase-1 activity assay rapidly monitors inflammasome 
      activation in cells.
PG  - 1-13
LID - S0022-1759(16)30275-7 [pii]
LID - 10.1016/j.jim.2017.03.004 [doi]
AB  - Inflammasomes are protein complexes induced by diverse inflammatory stimuli that 
      activate caspase-1, resulting in the processing and release of cytokines, IL-1beta 
      and IL-18, and pyroptosis, an immunogenic form of cell death. To provide a 
      homogeneous method for detecting caspase-1 activity, we developed a 
      bioluminescent, plate-based assay that combines a substrate, 
      Z-WEHD-aminoluciferin, with a thermostable luciferase in an optimized lytic 
      reagent added directly to cultured cells. Assay specificity for caspase-1 is 
      conferred by inclusion of a proteasome inhibitor in the lytic reagent and by use 
      of a caspase-1 inhibitor to confirm activity. This approach enables a specific 
      and rapid determination of caspase-1 activation. Caspase-1 activity is stable in 
      the reagent thereby providing assay convenience and flexibility. Using this assay 
      system, caspase-1 activation has been determined in THP-1 cells following 
      treatment with alpha-hemolysin, LPS, nigericin, gramicidin, MSU, R848, Pam3CSK4, and 
      flagellin. Caspase-1 activation has also been demonstrated in treated J774A.1 
      mouse macrophages, bone marrow-derived macrophages (BMDMs) from mice, as well as 
      in human primary monocytes. Caspase-1 activity was not detected in treated BMDMs 
      derived from Casp1(-/-) mice, further confirming the specificity of the assay. 
      Caspase-1 activity can be measured directly in cultured cells using the lytic 
      reagent, or caspase-1 activity released into medium can be monitored by assay of 
      transferred supernatant. The caspase-1 assay can be multiplexed with other assays 
      to monitor additional parameters from the same cells, such as IL-1beta release or 
      cell death. The caspase-1 assay in combination with a sensitive real-time monitor 
      of cell death allows one to accurately establish pyroptosis. This assay system 
      provides a rapid, convenient, and flexible method to specifically and 
      quantitatively monitor caspase-1 activation in cells in a plate-based format. 
      This will allow a more efficient and effective assessment of inflammasome 
      activation as well as enable high-throughput screening for inflammasome 
      modulators.
CI  - Copyright (c) 2017 The Authors. Published by Elsevier B.V. All rights reserved.
FAU - O'Brien, Martha
AU  - O'Brien M
AD  - Promega Corporation, 2800 Woods Hollow Rd, Madison, WI 53711, USA. Electronic 
      address: martha.obrien@promega.com.
FAU - Moehring, Danielle
AU  - Moehring D
AD  - Promega Corporation, 2800 Woods Hollow Rd, Madison, WI 53711, USA.
FAU - Munoz-Planillo, Raul
AU  - Munoz-Planillo R
AD  - Dept. of Pathology, University of Michigan Medical School, Ann Arbor, MI 48109, 
      USA.
FAU - Nunez, Gabriel
AU  - Nunez G
AD  - Dept. of Pathology, University of Michigan Medical School, Ann Arbor, MI 48109, 
      USA.
FAU - Callaway, Justin
AU  - Callaway J
AD  - Dept. of Microbiology and Immunology, School of Medicine, University of North 
      Carolina, Chapel Hill, NC 27599, USA.
FAU - Ting, Jenny
AU  - Ting J
AD  - Dept. of Microbiology and Immunology, School of Medicine, University of North 
      Carolina, Chapel Hill, NC 27599, USA.
FAU - Scurria, Mike
AU  - Scurria M
AD  - Promega Biosciences LLC, 277 Granada Dr, San Luis Obispo, CA 93401, USA.
FAU - Ugo, Tim
AU  - Ugo T
AD  - Promega Biosciences LLC, 277 Granada Dr, San Luis Obispo, CA 93401, USA.
FAU - Bernad, Laurent
AU  - Bernad L
AD  - Promega Biosciences LLC, 277 Granada Dr, San Luis Obispo, CA 93401, USA.
FAU - Cali, James
AU  - Cali J
AD  - Promega Corporation, 2800 Woods Hollow Rd, Madison, WI 53711, USA.
FAU - Lazar, Dan
AU  - Lazar D
AD  - Promega Corporation, 2800 Woods Hollow Rd, Madison, WI 53711, USA.
LA  - eng
PT  - Journal Article
DEP - 20170306
PL  - Netherlands
TA  - J Immunol Methods
JT  - Journal of immunological methods
JID - 1305440
RN  - 0 (Inflammasomes)
RN  - EC 1.13.12.- (Luciferases)
RN  - EC 3.4.22.36 (Caspase 1)
SB  - IM
MH  - Animals
MH  - Caspase 1/*metabolism
MH  - Cell Line
MH  - Enzyme Activation
MH  - Humans
MH  - Inflammasomes/*metabolism
MH  - Luciferases/metabolism
MH  - Luminescent Measurements/instrumentation/*methods
MH  - Macrophages/drug effects/metabolism
MH  - Mice
MH  - Monocytes/enzymology/*metabolism
MH  - Pyroptosis
MH  - Sensitivity and Specificity
OTO - NOTNLM
OT  - Bioluminescent
OT  - Caspase-1 assay
OT  - Inflammasome
OT  - Macrophages
OT  - Pyroptosis
OT  - THP-1 monocytes
EDAT- 2017/03/08 06:00
MHDA- 2017/09/16 06:00
CRDT- 2017/03/08 06:00
PHST- 2016/10/21 00:00 [received]
PHST- 2017/02/13 00:00 [revised]
PHST- 2017/03/03 00:00 [accepted]
PHST- 2017/03/08 06:00 [pubmed]
PHST- 2017/09/16 06:00 [medline]
PHST- 2017/03/08 06:00 [entrez]
AID - S0022-1759(16)30275-7 [pii]
AID - 10.1016/j.jim.2017.03.004 [doi]
PST - ppublish
SO  - J Immunol Methods. 2017 Aug;447:1-13. doi: 10.1016/j.jim.2017.03.004. Epub 2017 
      Mar 6.