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Human THP-1 Monocytes - ASC speck reporter cells
3-7 x 10e6 cells
Real-time monitoring of ASC-dependent inflammasomes
THP1-ASC-GFP cells are inflammasome reporter cells that allow the monitoring of ASC (apoptosis-associated speck-like protein with a CARD)-dependent inflammasome formation using fluorescence microscopy.
These cells are derived from the THP-1 human monocytic cell line and stably express a gene encoding an ASC::GFP fusion protein for which expression is driven by an NF-κB-inducible promoter. Hence, in resting cells, no GFP signal is detected.
Upon the first step of inflammasome activation (‘priming”), NF-kB-dependent ASC::GFP expression is induced and can be observed throughout the cytoplasm. Following the second step of inflammasome activation, ASC::GFP polymerizes to form a macromolecular, micrometer-sized complex.
The presence of ASC::GFP positive cells and localization of fluorescent ASC specks can be determined by using time-lapse confocal or high-resolution fluorescence microscopy.
The number of ASC specks is dependent on the inflammasome inducer used; Nigericine promotes the formation of numerous specks, whereas ATP induction leads to the accumulation of fewer specks . In most cells only one speck forms upon inflammasome activation [1,2].
THP1-ASC-GFP cells are resistant to Zeocin™.
1. Hoss F. et al., 2017. Assembly and regulation of ASC specks. Cell. Mol. Life Sci. 74:1211-29.
2. Stutz A. et al., 2013. ASC speck formation as a readout for inflammasome activation. Methods Mol Biol. 1040:91-101.
3. Zha Q. et al., 2016. ATP-induced inflammasome activation and pyroptosis is regulated by AMP-activated protein kinase in macrophages. Front Immunol. 7:597
THP1-ASC-GFP cells were primed with 1 μg/ml LPS-EK for 3 hours, inducing the expression of the ASC::GFP fusion protein in the cytoplasm (middle panel). Cells were then incubated with 250 ng/ml complexed Poly(dA:dT) and ASC speck formation was monitored over 1 to 3 hours post-activation. In most cells, only one speck forms upon inflammasome activation (arrows in right pannel). Scale bar: 50 μm.
Antibiotic resistance: Zeocin
Growth medium: RPMI 1640, 2 mM L-glutamine, 25 mM HEPES, 10% heat-inactivated fetal bovine serum, 100 μg/ml Normocin™, Pen-Strep (50 U/ml-50 μg/ml)
- The functionality of THP1 ASC-GFP cells has been tested using inflammasome inducers, such as the microbial toxin nigericin and transfected poly(dA:dT).
- The stability of this cell line for 20 passages following thawing has been verified.
- THP1 ASC-GFP cells are guaranteed mycoplasma-free.
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- 1 vial of THP1 ASC-GFP cells (3-7 x 106 cells) in Freezing Medium.
- 1 ml Normocin™ (50 mg/ml). Normocin™ is a formulation of three antibiotics active against mycoplasmas, bacteria and fungi.
- 100 μl Zeocin (100 mg/ml).
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