α ripk1

Manufactured by BD

α-RIPK1 is a lab equipment product designed for use in research applications. It is a protein that plays a key role in the regulation of programmed cell death pathways. The core function of α-RIPK1 is to serve as a tool for researchers studying cell signaling mechanisms and cell death processes.

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Lab products found in correlation

α actin, by Santa Cruz Biotechnology (2 mentions) α myc, by Merck Group (2 mentions) α casp8, by Santa Cruz Biotechnology (2 mentions) α fadd, by Santa Cruz Biotechnology (2 mentions) Nec 1, by Merck Group (2 mentions) Pcdna3, by Thermo Fisher Scientific (1 mentions) Tumor necrosis factor (tnf), by Enzo Life Sciences (1 mentions) α flag m2, by Merck Group (1 mentions) α p jnk, by Cell Signaling Technology (1 mentions) α tubulin, by Merck Group (1 mentions) α pp38, by Cell Signaling Technology (1 mentions) α erk, by Cell Signaling Technology (1 mentions) α perk, by Cell Signaling Technology (1 mentions) α p p65, by Cell Signaling Technology (1 mentions) α caspase 8, by Cell Signaling Technology (1 mentions) α sharpin, by Proteintech (1 mentions) α cd69 pe, by Thermo Fisher Scientific (1 mentions) α cd3 apc, by Thermo Fisher Scientific (1 mentions) Z vad fmk, by Apexbio (1 mentions) α ripk3, by Novus Biologicals (1 mentions)

Close spelling variants of this product

RIPK1 (31 citations)

4 protocols using α ripk1

Cloning and Mutagenesis of Constructs

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Constructs were generated by PCR and cloned into pAc, pMT, pcDNA3 (Invitrogen) or pEF6 and verified by sequencing. Specific point mutations were generated using site-directed mutagenesis with Pfu Turbo polymerase (Stratagene). The Ub-sgg10 fusion construct was generated by PCR using Easy-A polymerase and cloned into pMT-V5/His. The following antibodies were used: α-MYC (Sigma, M5546, 1:2,000), α-HA (Roche, 11867423001, 1:2,000), α-V5 (Serotec, MCA1360, 1:2,000), α-RIPK1 (BD Biosciences, 610459,1:1,000 for western blotting (WB) and 1:50 for immunofluorescence studies), α-Actin (Santa Cruz, sc-1615, 1:4,000), α-MYO7A (Developmental Studies Hybridoma Bank, 138-1-s, 1:1,000 for WB and 1:50 for immunofluorescence studies), α-CASP8 for WB (MBL, M032-3, 1:5,000), α-CASP8 to detect cleaved CASP8 (for human: R&D, AF1650, 1:2,000; for mouse: Cell Signaling Technology, 9429, 1:1,000), rabbit and goat α-CASP8 (Santa Cruz Biotechnology, sc-7890 and sc-6136, both 1:50 for immunofluorescence studies), CF488A-donkey α-mouse IgG (Biotium, A21202, 1:1,000), CF633-donkey α-rabbit IgG (Biotium, 20125, 1:1,000) and CF633-donkey α-goat IgG (Biotium, 20127, 1:1,000). SM (SM164) was a gift from Shaomeng Wang (University of Michigan) and TNF was obtained from Enzo Life Sciences. Uncropped WBs are shown in Supplementary Fig. 5.

Orme M.H., Liccardi G., Moderau N., Feltham R., Wicky-John S., Tenev T., Aram L., Wilson R., Bianchi K., Morris O., Monteiro Domingues C., Robertson D., Tare M., Wepf A., Williams D., Bergmann A., Gstaiger M., Arama E., Ribeiro P.S, & Meier P. (2016). The unconventional myosin CRINKLED and its mammalian orthologue MYO7A regulate caspases in their signalling roles. Nature Communications, 7, 10972.

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RIPK1 Kinase Inhibitor Mechanism Study

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The GSK’963 RIPK1 kinase inhibitor was provided by GSK. The following antibodies were used: α-RIPK1 (BD Biosciences, 610459), α-HOIL (gift from Henning Walczak), α-cIAP1 (Enzo, ALX-803-335-C100), α-TNFR1 (Abcam, 19139), α-Actin (Santa Cruz Biotechnology, sc-1615), α-P-p65 (Cell Signaling, 3033), α-p65 (Cell Signaling, 8242), α-IkBα (Santa Cruz, sc-371), α-P-p38 (Cell Signaling, 9215), α-p38 (Cell Signaling, 9212), α-P-JNK (Cell Signaling, 9255), α-JNK (Santa Cruz Biotechnology, sc-571), α-P-ERK (Cell Signaling, 9101), α-ERK (gift from Chris Marshall) α-caspase-8 (Cell Signaling, 9429), α-FLAG [M2] (SIGMA, M8823), α-Ub (Dako, Z0458), α-FLIP (Adipogene, AG-20B-0056), α-FADD (Santa Cruz Biotechnology, sc-6036), α-RIPK3 (ProSci, 2283), α-Tubulin (SIGMA, T9026), α-SHARPIN (Proteintech, 14626-1-AP), α-TRAF2 (Cell Signaling, 4712), α-CD8-PE-Cy7, GR-1-PE-Cy7, CD11c-FITC, CD4-FITC, CD11b-Cy5, B220-FITC (gift from Henning Walczak), α-CD69-PE (eBioscience, 12-0691-82), α-CD3-APC (eBioscience, 47-0032-82), and α-CD16 (eBioscience, 14-0161-82).

Annibaldi A., Wicky John S., Vanden Berghe T., Swatek K.N., Ruan J., Liccardi G., Bianchi K., Elliott P.R., Choi S.M., Van Coillie S., Bertin J., Wu H., Komander D., Vandenabeele P., Silke J, & Meier P. (2018). Ubiquitin-Mediated Regulation of RIPK1 Kinase Activity Independent of IKK and MK2. Molecular Cell, 69(4), 566-580.e5.

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Immunoprecipitation of RIPK1 and ZBP1 from BMDMs

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Indicated BMDMs were plated on 6-well tissue-culture-treated plates, stimulated as indicated, and harvested in immunoprecipitation lysis buffer (0.5% Triton X, 50 mM Tris Base [pH 7.4], 150 mM NaCl, 2 mM ethylenediaminetetraacetic acid, 2 mM ethylene glycol tetraacetic acid, 1X protease inhibitor mixture). Lysed cells were rotated for 60 min at 4 °C with intermittent vortexin and centrifuged at 5,000 X g for 5 min, and the supernatant was incubated with α-RIPK1 (BD Biosciences 610459) or α-ZBP1 (AG-20B-0010-C100) antibody-conjugated protein G agarose beads (Cell Signaling Technology 37478). Samples were washed three times in immunoprecipitation lysis buffer, and protein complexes were eluted with 1X Laemmli buffer containing 5% β-mercaptoethanol at 90 °C for 15 min.

Muendlein H.I., Connolly W.M., Magri Z., Jetton D., Smirnova I., Degterev A., Balachandran S, & Poltorak A. (2022). ZBP1 promotes inflammatory responses downstream of TLR3/TLR4 via timely delivery of RIPK1 to TRIF. Proceedings of the National Academy of Sciences of the United States of America, 119(24), e2113872119.

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Investigating Cell Death Pathways

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The following reagents were used: zVAD-FMK (10 μM, Apex Bio), QVD (10 μM, Apex Bio), SM-164 (100 nM, gift from Shaomeng Wang), GSK’963 (referred as RIPK1i, 100 nM, GlaxoSmithKline), Nec-1 (10 μM Merck), Nec-1s (10 μM Merck), RO-3306 (9 μM, Merck), Thymidine (Sigma), BIM peptide (Kind gift of Tony Letai), FLAG-hTNF (10 ng/ml, Enzo), MG132 (1-20 μM, as indicated per cell line, see below, SIGMA), Recombinant Casp8 (1U Enzo). The following antibodies were used for western blotting: α-RIPK1 (1:1000, BD Biosciences), α-HA (1:1000, Roche), α-PLK1 (1:1000, Bethyl Laboratories), α-PLK1-pT210 (1:2000, AbCam), α-Cyclin B (1:1000, Cell Signaling), α-BUBR1 (1:1000, BD Bioscience), α-BUBR1-pT680 (1:1000, Kind gift of Geert J.P.L Kops), α-BUBR1-pT676 (1:1000, Kind gift of Erich A. Nigg), α-pH-H3 (1:2000, Millipore), α-Casp8 (for WB - post IP, 1:5000, MBL), α-Casp8 - for IP (7.5 μg/ml, C-20, Santa Cruz Biotechnology, α-FADD – for IP (7.5 μg/ml, Santa Cruz), α-FADD (1:1000 BD Biosciences), α-RIPK3 (1:1000 Proscience), α-RIPK3 (1:1000 Novus Biological) α-cFLIP (1:1000, Enzo), α-Myc (1:1000, clone 9E10,SIGMA), α-HSP90 (1:1000 Santa Cruz Biotechnology).

Liccardi G., Ramos Garcia L., Tenev T., Annibaldi A., Legrand A.J., Robertson D., Feltham R., Anderton H., Darding M., Peltzer N., Dannappel M., Schünke H., Fava L.L., Haschka M.D., Glatter T., Nesvizhskii A., Schmidt A., Harris P.A., Bertin J., Gough P.J., Villunger A., Silke J., Pasparakis M., Bianchi K, & Meier P. (2019). RIPK1 and Caspase-8 Ensure Chromosome Stability Independently of Their Role in Cell Death and Inflammation. Molecular Cell, 73(3), 413-428.e7.

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