Necrostatin-1

For research use only.

Catalog No.S8037 Synonyms: Nec-1

91 publications

Necrostatin-1 Chemical Structure

CAS No. 4311-88-0

Necrostatin-1 (Nec-1) is a specific RIP1 (RIPK1) inhibitor and inhibits TNF-α-induced necroptosis with EC50 of 490 nM in 293T cells. Necrostatin-1 also blocks IDO and suppresses autophagy and apoptosis.

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Selleck's Necrostatin-1 has been cited by 91 publications

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Choose Selective RIP kinase Inhibitors

Biological Activity

Description Necrostatin-1 (Nec-1) is a specific RIP1 (RIPK1) inhibitor and inhibits TNF-α-induced necroptosis with EC50 of 490 nM in 293T cells. Necrostatin-1 also blocks IDO and suppresses autophagy and apoptosis.
Features A powerful tool for characterizing the role of necroptosis with characterized primary target.
Targets
RIP1 [1]
(293T cells)
490 nM(EC50)
In vitro

Necrostatin-1 (1-100 μM) inhibits the autophosphorylation of overexpressed and endogenous RIP1.It is found RIP1 is the primary cellular target responsible for the antinecroptosis activity of Necrostatin-1. [1] Necrostatin-1 efficiently suppresses necroptotic cell death triggered by an array of stimuli in a variety of cell types. Necrostatin-1, previously identified as small-molecule inhibitor of necroptosis, inhibits RIP kinase-induced necroptosis and inhibits TNF-α-induced necroptosis in jurkat cells with EC50 of 490 nM. [2]

Cell Data
Cell Lines Assay Type Concentration Incubation Time Formulation Activity Description PMID
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Jurkat  M1z3XWZ2dmO2aX;uJGF{e2G7 NH7RRmMzODEEoN88cS=> NFK0SmQ{OCCvaX6= NWnkSpVsemWmdXPld:KhVmGnZ3zldoliKG[xd3zldokucW6mdXPl[EBz\WGldHn2[UBwgHmpZX6gd5Bm[2mnczCoVm9UMSCpZX7ldoF1cW:w MofiQIEhfGG{Z3X0QUdg[myjbnunJIhz\WZ;J3j0eJB{Qi9xcIXicYVlNm6lYnmucoxuNm6raD7nc5YwOjF3M{WwNlAoRjJzNUO1NFIxRC:jPh?=
Jurkat  NIfNXohEgXSxdH;4bYNqfHliQYPzZZk> MlXaOVAwKDFyMD:yNFDDqM7:bR?= NWroeIs5OS9|IHi= MX3y[YR2[2W|wrDOZYVodGW{aXGg[o94dGW{aT3pcoR2[2WmIHP5eI91d3irY3n0fS=> MkPUQIEhfGG{Z3X0QUdg[myjbnunJIhz\WZ;J3j0eJB{Qi9xcIXicYVlNm6lYnmucoxuNm6raD7nc5YwOjF3M{WwNlAoRjJzNUO1NFIxRC:jPh?=
SW13 M1v2bWNmdGxiVnnhZoltcXS7IFHzd4F6 MWWxNFAh|ryP NEnndo4zPCCq NHnEU5JFVVOR MWrpcoNz\WG|ZYOgZ4VtdHWuYYKgd5Vzfmm4YXy= NFfqVIw9[SC2YYLn[ZQ:L1:kbHHub{chcHKnZk2nbJR1eHN8Lz;weYJu\WRwbnPibU5vdG1wbnnoModwfi9{MkGzOlgyQCd-MkKxN|Y5OTh:L3G+
8505c MWjD[YxtKF[rYXLpcIl1gSCDc4PhfS=> MWKxNFAh|ryP MmfTNlQhcA>? M3;md2ROW09? NHzIfG5qdmO{ZXHz[ZMh[2WubIXsZZIhe3W{dnn2ZYw> MlHTQIEhfGG{Z3X0QUdg[myjbnunJIhz\WZ;J3j0eJB{Qi9xcIXicYVlNm6lYnmucoxuNm6raD7nc5YwOjJzM{[4NVgoRjJ{MUO2PFE5RC:jPh?=
TPC-1 MlXvR4VtdCCYaXHibYxqfHliQYPzZZk> MoPmNVAxKM7:TR?= M3fqbVI1KGh? MVTEUXNQ Ml\abY5kemWjc3XzJINmdGy3bHHyJJN2en[rdnHs NELGUW49[SC2YYLn[ZQ:L1:kbHHub{chcHKnZk2nbJR1eHN8Lz;weYJu\WRwbnPibU5vdG1wbnnoModwfi9{MkGzOlgyQCd-MkKxN|Y5OTh:L3G+
L929sA M{TvSWFxd3C2b4Ppd{BCe3OjeR?= M3HSWFExKM7:TR?= NFTU[WgyKGh? M3nrWIFjem:pYYTld{B1cGViaX70[ZJi[3Srb36gc4Yh[2G|cHHz[U05KHerdHigSmFFTA>? NXTxSG9rRGFidHHy[4V1RSehYnzhcosoKGi{ZX[9K4h1fHC|Oj:vdJVjdWWmLn7jZokvdmyvLn7pbE5od3ZxMkKzOlI4PjdpPkKyN|YzPzZ5PD;hQi=>
L929sA MnWzRZBweHSxc3nzJGF{e2G7 MkD4NVAh|ryP Mne5NUBp MknxdoV{[3WnczDj[YxteyCneIDy[ZN{cW6pIGLJVGsy|pSLRDDmdo9uKFSQRj3pcoR2[2WmIHHwc5B1d3Orcx?= MWW8ZUB1[XKpZYS9K39jdGGwazegbJJm\j1paIT0dJM7Ny:ydXLt[YQvdmOkaT7ucI0vdmmqLnfvek8zOjN4Mke2O{c,OjJ|NkK3Olc9N2F-
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K562 NELTRVVHfW6ldHnvckBCe3OjeR?= MmXoOlAh|ryP M1i4XVEzKGh? NFnBXXpifWevZX70d{B1cGViY3HzdIF{\S1|IHHjeIl3cXS7 MkjyQIEhfGG{Z3X0QUdg[myjbnunJIhz\WZ;J3j0eJB{Qi9xcIXicYVlNm6lYnmucoxuNm6raD7nc5YwOjJ6M{e2PFkoRjJ{OEO3Olg6RC:jPh?=
HL60/Adr M4f0dGZ2dmO2aX;uJGF{e2G7 NWC4dWNSPjBizszN NGD0[ngyOiCq MXTheYdu\W62czD0bIUh[2G|cHHz[U0{KGGldHn2bZR6 NYjTWmNFRGFidHHy[4V1RSehYnzhcosoKGi{ZX[9K4h1fHC|Oj:vdJVjdWWmLn7jZokvdmyvLn7pbE5od3ZxMkK4N|c3QDlpPkKyPFM4Pjh7PD;hQi=>
HL60 Mnj1SpVv[3Srb36gRZN{[Xl? M3n2S|YxKM7:TR?= NHLocWUyOiCq Mn7mZZVodWWwdIOgeIhmKGOjc4Dhd4UuOyCjY4Tpeol1gQ>? M3PUT|xiKHSjcnfleF0oZ2KuYX7rK{BpemWoPTfoeJRxezpxL4D1Zo1m\C6wY3LpMo5tdS6waXiu[493NzJ{OEO3Olg6Lz5{MkizO|Y5QTxxYU6=
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HL60 NFjwWWNCeG:ydH;zbZMhSXO|YYm= NG\4Xm03OCEQvF2= NVHHWHJrOTJiaB?= MVTlcohidmOnczDzbIlsd26rbj3pcoR2[2WmIHHwc5B1d3Orcx?= M3LISlxiKHSjcnfleF0oZ2KuYX7rK{BpemWoPTfoeJRxezpxL4D1Zo1m\C6wY3LpMo5tdS6waXiu[493NzJ{OEO3Olg6Lz5{MkizO|Y5QTxxYU6=
SH-EP NVPTR4ZySXCxcITvd4l{KEG|c3H5 MWKxNQKBkc7:TdMg M{P0Olcz6oDLaNMg NWjadVhycW6qaXLpeJMhUUGSIHnubIljcXSxcj2gZY5lKEyneHH0eY12dWGkLXnu[JVk\WRiYYDvdJRwe2m| MXG8ZUB1[XKpZYS9K39jdGGwazegbJJm\j1paIT0dJM7Ny:ydXLt[YQvdmOkaT7ucI0vdmmqLnfvek8zOjh7MEOyNkc,OjJ6OUCzNlI9N2F-
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ΔN-Karpas 299  MnvXR5l1d3SxeHnjbZR6KEG|c3H5 NFXKVlAzOMLizszN MofXNVYhcA>? MkfZbY5pcWKrdIOgR2Q{OC2rbnT1Z4VlKGOnbHyg[IVifGh? MmDuQIEhfGG{Z3X0QUdg[myjbnunJIhz\WZ;J3j0eJB{Qi9xcIXicYVlNm6lYnmucoxuNm6raD7nc5YwOjN3NEW5N|goRjJ|NUS1PVM5RC:jPh?=
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... Click to View More Cell Line Experimental Data

Assay
Methods Test Index PMID
Immunofluorescence
RIP1 / RIP3 ; 

PubMed: 30462730     


Immunofluorscence data showed an increased expression of RIP1 and RIP3 in Pg-LPS group and an eminent reduction of RIP1 in the Pg-LPS+Nec-1 group.

30462730

Protocol

Kinase Assay:

[1]

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RIP1 kinase assay:

Phosphorylation of RIP1 requires its kinase activity. Expression constructs of FLAGtagged wild-type (WT) or a kinase-inactive pointmutant of RIP1 (K45M) are are transfected into 293T cells and RIP1 kinase assay is performed as described in the Methods in the presence of [γ-32P]ATP for 30 min at 30℃. Samples are subjected to SDS-PAGE and RIP1 band is visualized by autoradiography. Relative intensities of radioactive bands are quantified and are shown (ratio) in this and all other autoradiographs. In parallel to kinase reactions, a sample of beads is subjected to western blot analysis using anti-RIP1 antibody to ensure equal protein amounts in kinase reactions.
Cell Research:

[2]

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  • Cell lines: Jurkat, BALB/c 3T3, SV40-transformed MEF, L929
  • Concentrations: 0.01-100 μM
  • Incubation Time: --
  • Method:

    Cells are seeded in 96-well plates (white plates for luminescent assays; black plates for fluorescent assays; clear plates for MTT assay) at the density of 5,000-10,000 cells per well for adherent cells or 20,000-50,000 cells per well for suspension cells in 100 μl of the appropriate phenol red-free media. After incubation, we determined cell viability using one of the following methods. For the ATP assay, we used luminescence-based commercial kits and analyzed luminescence using a Wallac Victor II plate reader. For Sytox assay, we incubated cells with 1 μM Sytox Green reagent for 30 min at 37℃, and then performed fluorescent reading. Subsequently, we added 5 μl of 20% Triton X-100 solution into each well to produce maximal lysis and incubated cells for 1 h at 37℃, then performed the second reading. We calculated the ratio of values before and after Triton treatment and normalized it to the relevant controls not subjected to cytotoxic stimuli, as indicated in figure legends. For the MTT assay, we used the CellTiter 96 AQueous Non-Radioactive Cell Proliferation Assay kit. For PI exclusion assays, we added 2 μg/ml PI into the medium and immediately analyzed samples using FACSCalibur. For PI-annexin V assay we used the ApoAlert Annexin V-EGFP Apoptosis Kit. For DioC6 staining, we incubated cells with 40 nM DiOC6 for 30 min at 37 ℃, washed once and analyzed in FACSCalibur. For ROS analysis, we incubated cells with 5 μM dihydroethidium for 30 min at 37 ℃, washed once and analyzed in FACSCalibur. EM analyses are performed at the Harvard Medical School EM facility. We acquired bright-field images of the cells using an Axiovert 200 microscope.


    (Only for Reference)

Solubility (25°C)

In vitro DMSO 51 mg/mL warmed (196.66 mM)
Water Insoluble
Ethanol Insoluble
In vivo Add solvents to the product individually and in order(Data is from Selleck tests instead of citations):
5% DMSO+45% PEG 300+ddH2O
For best results, use promptly after mixing.
10mg/mL

* Please note that Selleck tests the solubility of all compounds in-house, and the actual solubility may differ slightly from published values. This is normal and is due to slight batch-to-batch variations.

Chemical Information

Molecular Weight 259.33
Formula

C13H13N3OS

CAS No. 4311-88-0
Storage powder
in solvent
Synonyms Nec-1
Smiles CN1C(=O)C(NC1=S)CC2=CNC3=CC=CC=C32

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Tech Support

Answers to questions you may have can be found in the inhibitor handling instructions. Topics include how to prepare stock solutions, how to store inhibitors, and issues that need special attention for cell-based assays and animal experiments.

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Cell Lines Assay Type Concentration Incubation Time Formulation Activity Description PMID