Anti ulk1

Manufactured by Santa Cruz Biotechnology

Sourced in United States

Anti-ULK1 is a laboratory reagent that is used to detect and quantify the expression of the ULK1 protein, which is a key regulator of autophagy. It is a specific antibody that can be used in various immunoassay techniques, such as Western blotting, immunohistochemistry, and immunoprecipitation, to analyze the presence and levels of ULK1 in biological samples.

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Lab products found in correlation

Anti beclin 1, by Cell Signaling Technology (3 mentions) Anti gfp, by Santa Cruz Biotechnology (2 mentions) Anti myc, by Cell Signaling Technology (1 mentions) Anti cleaved caspase 3, by Cell Signaling Technology (1 mentions) Anti β actin peroxidase conjugated antibody, by Merck Group (1 mentions) Anti cleaved caspase 7, by Cell Signaling Technology (1 mentions) Anti cleaved parp, by Cell Signaling Technology (1 mentions) Anti h3k4me3, by Merck Group (1 mentions) Anti atg5, by Merck Group (1 mentions) Anti h3k27me3, by Merck Group (1 mentions) Anti olig2, by Merck Group (1 mentions) Anti gfap, by BD (1 mentions) Anti sox2, by Cell Signaling Technology (1 mentions) Anti atg7, by Cell Signaling Technology (1 mentions) Anti rna pol 2, by Active Motif (1 mentions) Anti beclin 1, by BD (1 mentions) Anti tubulin beta 3 isoform, by Merck Group (1 mentions) Anti g9a, by Abcam (1 mentions) Anti h3k9me2, by Abcam (1 mentions) Anti lc3, by Merck Group (1 mentions)

Spelling variants of this product

Anti-ULK1 antibody (3 citations) Rabbit anti-ULK1 (2 citations)

9 protocols using anti ulk1

In Vitro p38α Kinase Assay

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In vitro p38α kinase assay was performed following the manufacturer’s instructions. In brief, ULK1 proteins were immunoprecipitated from cells with either anti-MYC (Cell Signaling Technology) or anti-ULK1 (Santa Cruz Biotechnology, Inc.) antibodies. Then, the immunoprecipitation (IP) products were washed three times with IP buffer (10% glycerol, 50 mM Tris, pH 7.7, 150 mM NaCl, 0.5% NP-40, 1× DTT, 1× protease inhibitors, and 0.1 µM okadaic acid) followed by 1× kinase buffer (25 mM Tris-HCl, pH 7.5, 5 mM β-glycerophosphate, 2 mM DTT, 0.1 mM Na₃VO₄, and 10 mM MgCl₂). The immunoprecipitated products were incubated with 100 ng p38α MAPK in kinase buffer containing 25 µM cold ATP with or without 2 µCi [γ-³²P]ATP under 30°C for 30 min, and the reaction was terminated by adding SDS sample buffer, heated at 95°C for 5 min, and then subjected to SDS-PAGE and autoradiography.

He Y., She H., Zhang T., Xu H., Cheng L., Yepes M., Zhao Y, & Mao Z. (2018). p38 MAPK inhibits autophagy and promotes microglial inflammatory responses by phosphorylating ULK1. The Journal of Cell Biology, 217(1), 315-328.

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Autophagy Regulation in Stem Cells

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Reagent and antibody sources were as follows: acridine orange, ATRA (all-trans-retinoic acid), bafilomycin A1 (BafA1), BIX01294 trihydrochloride hydrate, DAPI (4′,6-diamidino-2-phenylindole dihydrochloride), laminin, 3-methyladenine (3MA), MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide), anti-LC3 (catalog no. L7543) and anti-β-Actin-peroxidase conjugated antibody (catalog no. A3854) (Sigma-Aldrich, Munich, Germany), anti-ATG5 (catalog no. 2630), anti-ATG7 (catalog no. 2631), anti-cleaved caspase 3 (catalog no. 9661), anti-cleaved caspase 7 (catalog no. 9491), anti-cleaved PARP (poly (ADP-ribose) polymerase-1) (catalog no. 9541), anti-SOX2 (catalog no. 3579) (Cell Signaling Technology, Beverly MA, USA), anti-ULK1 (catalog no. sc-33182) (Santa Cruz Biotechnology, Dallas, Texas, USA), anti-Beclin1 (catalog no. 612112), anti-GFAP (catalog no. 556330) (BD Pharmingen San Jose, CA, USA), anti-H3K4me3 (catalog no. 07-473), anti-H3K27me3 (catalog no. ABE44), anti-OLIG2 (catalog no. AB9610), anti-Tubulin beta III isoform (catalog no. MAB1637) (Millipore, Temecula, CA, USA), anti-H3K9me2 (catalog no. ab1220), anti-G9a (catalog no. ab40542) (Abcam, Cambridge, UK), anti-RNA Pol II (catalog no. 39097) (Active Motif, Carlsbad, CA, USA) and anti-NESTIN (catalog no. MAB1259) (R&D Systems, Minneapolis, MN, USA).

Ciechomska I.A., Przanowski P., Jackl J., Wojtas B, & Kaminska B. (2016). BIX01294, an inhibitor of histone methyltransferase, induces autophagy-dependent differentiation of glioma stem-like cells. Scientific Reports, 6, 38723.

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Antibody Targeting of Lon Protease Autophagy Pathway

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Antibodies to human Lon were produced as described previously [35 (link)]. The following primary antibodies were used in this study: Autophagy Induction Antibody Sampler Kit containing Anti-ULK1 monoclonal antibody, anti-p-ULK1 S555, anti-ATG13, anti-p-ATG13 S355, anti-FIP200, anti-ATG101. Anti-NFkB anti-LC3B, anti-LAMP1, anti-VDAC, anti-Calnexin, anti-Tubulin, and ULK1 antibody sampler Kit containing anti-ATG14, anti-p-ATG14 S29, anti-Beclin1, anti-p-Beclin1 S15, were purchased from Cell signaling technology. Anti-FACL4, anti-GAPDH, and anti-β-actin were purchased from Gentex also the Gentex helped in producing in-house antibody anti-FUNDC1 and anti-p-FUNDC1 S17 which were raised in rabbit and purified. Anti-HSP60 and anti-Aconitase2 from santa cruz. Anti-myc and and anti-Flag were obtained from Merck Millipore. Anti-HIF1α was obtained from Life science bio. Anti-ULK1 for immuofluorescence studies was obtained from Santa cruz. Cobalt chloride (CoCl₂), SBI-0206965, Bafilomycin A1 and N-Acetyl Cysteine (NAC) were purchased from sigma, dissolved in DMSO and stored at −20 °C.

Ponneri Babuharisankar A., Kuo C.L., Chou H.Y., Tangeda V., Fan C.C., Chen C.H., Kao Y.H, & Lee A.Y. (2023). Mitochondrial Lon-induced mitophagy benefits hypoxic resistance via Ca2+-dependent FUNDC1 phosphorylation at the ER-mitochondria interface. Cell Death & Disease, 14(3), 199.

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Immunoblotting Antibody Validation Protocol

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The following primary antibodies were used: HRP-conjugated anti-GST (GE Healthcare, RPN1236); anti-MYC (CRUK, 9E10; Abcam, ab9106); anti-GFP (CRUK, 3E1; Santa Cruz, sc-8334); anti-GABARAP (Abgent, AP1821a); anti-actin (Abcam, ab8227), anti-FLAG M2 (SIGMA, F1804), anti-TGN46 (Serotec, AHP500), anti-beta-tubulin (Abcam, ab6046), anti-ULK1 (Santa Cruz, sc-33182); anti-ATG13⁶⁵ (link); anti-FIP200 (Bethyl Labs, A301-536A-1); anti-FEZ2 (SIGMA, HPA0355978). Secondary antibodies for IF were: anti-rabbit IgG Alexa Fluor 555 and 647; anti-mouse IgG Alexa Fluor 555, 647; anti-sheep IgG Alexa Fluor 647 (all from Life Technologies). HRP-conjugated secondary antibodies used for WB were from GE Healthcare.

Wirth M., Mouilleron S., Zhang W., Sjøttem E., Princely Abudu Y., Jain A., Lauritz Olsvik H., Bruun J.A., Razi M., Jefferies H.B., Lee R., Joshi D., O'Reilly N., Johansen T, & Tooze S.A. (2021). Phosphorylation of the LIR Domain of SCOC Modulates ATG8 Binding Affinity and Specificity. Journal of Molecular Biology, 433(13), 166987.

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Mitotic Checkpoint Protein Localization

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The antibodies used were anti-MAD1 (proteintech, #18322-1-AP; Santa Cruz, #sc-47746), anti-CENP-B (Santa Cruz, #sc-376283; #sc-22788), anti CDC20 (Santa Cruz, #sc-5296), anti-MAD2 (proteintech, #10337-1-AP; Covance, #PRB-452C-200), anti-BUBR1 (proteintech, #11504-2-AP), anti-cyclinB1 (abclonal, #A2056), anti-ZW10 (abcam, #ab21582), anti-Flag-tag (Sigma-Aldrich, #F1804), anti-ULK1 (Santa Cruz, #sc-33182; proteintech, #20986-1-AP), anti-Atg13 (CST, #13468S), anti-FIP200 (proteintech, #17250-1-AP), anti-Atg3 (MBL, #M133-3), anti-GFP-tag (Santa Cruz, #sc-9996), anti-His-tag (MBL, #D291-3), anti-BUB1 (proteintech, #13330-1-AP), anti-BUB3 (proteintech, #27073-1-AP), anti-H3S10 (abcam, #ab32017), anti-Knl1 (abclonal, A13108), anti-ROD (abclonal, A13064), anti-ZWILCH (proteintech, #14281-1-AP), anti-α-tubulin (CST, #3873T) and anti-actin (Santa Cruz, #sc-7210). A phospho (ph)-MAD1-S546 specific antibody was generated by Beijing Biodragon Immunotechnologies Co. Ltd.
Treatments included thymidine (Sigma, T9250), RO3306 (Selleck, S7747), Taxol (Santa Cruz, sc-201439), nocodazole (Selleck, S2775), vinblastine (Selleck, S4505).

Yuan F., Jin X., Li D., Song Y., Zhang N., Yang X., Wang L., Zhu W.G., Tian C, & Zhao Y. (2019). ULK1 phosphorylates Mad1 to regulate spindle assembly checkpoint. Nucleic Acids Research, 47(15), 8096-8110.

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Protein Extraction and Western Blot Analysis

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Protein was extracted from goat LD tissues and primary myoblast cells using a Total Histone Extraction Kit (Beyotime Biotech Co., Ltd., Shanghai, China); protein concentrations were determined using a BCA protein assay kit (Beyotime). The 30 µg of protein per sample was resolved using 12% SDS–PAGE and then transferred onto PVD membranes activated with methanol. After transfer, membranes were blocked with blocking buffer (Beyotime) for 6 h at 4 °C, and then incubated with primary antibodies overnight at 4 °C. The primary antibodies were anti-ULK1 (1:100) (Santa Cruz Biotechnology, New York, NY, USA), anti-VMP1 (1:500), anti-beclin-1 (1:1000), anti-caspase-3 (1:1000), anti-mTOR mammalian (1:1000), and anti-GAPDH (1:1000) (all from Cell Signaling Technology, New York, NY, USA). After incubation, the membranes were rinsed with wash buffer (Beyotime) and incubated for 2 h at 4 °C with secondary antibodies. The secondary Ab was HRP-conjugated goat anti-rabbit IgG (H + L) (Proteintech, Wuhan, China), used at dilutions recommended by Beyotime.

Liu Y., Zhou Z., Li K., Wang P., Chen Y., Deng S., Li W., Yu K, & Wang K. (2022). VMP1 Regulated by chi-miR-124a Effects Goat Myoblast Proliferation, Autophagy, and Apoptosis through the PI3K/ULK1/mTOR Signaling Pathway. Cells, 11(14), 2227.

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Immunoblotting Techniques for Protein Analysis

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The following antibodies were used: anti-TG2 (Thermo Scientific, CUB7402), anti-PKM2 (Cell Signalling, D78A4), anti-EEF1A1 (Milipore, 05-235), anti-Hsc70 (Abcam, 51052), HSPA1A (Santa Cruz, 7947), anti-Biotin (Abcam, 1227), anti-p62/SQSTM1 (mbl, PM045), anti-LC3 (Novus Biologicals, NB100-2331), anti-GAPDH (Sigma, G9545), anti-Actin (Sigma, 2066), anti-Beclin1 (Santa Cruz, 10086), anti-HIF1-beta (Novus Biologicals, NB100-124), anti-ULK1(Santa Cruz, 33182), anti-phospho-tyrosine (Cell Signaling, 9411) and HRP-conjugated secondary antibodies (Bio-Rad Laboratories).

Altuntas S., Rossin F., Marsella C., D'Eletto M., Hidalgo L.D., Farrace M.G., Campanella M., Antonioli M., Fimia G.M, & Piacentini M. (2015). The transglutaminase type 2 and pyruvate kinase isoenzyme M2 interplay in autophagy regulation. Oncotarget, 6(42), 44941-44954.

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Comprehensive Antibody Validation Protocol

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Anti-BRUCE (BD, 611193; 1:500), anti-Ubiquitin (P4D1, Santa Cruz, sc-8017; 1:1,000), anti-Myc (9E10, Covance, MMS-150P; 1:1,000), anti-GFP (Santa Cruz, sc-9996; 1:1,000), anti-mCherry (Clontech, 632543; 1:1,000), anti-Alpha-Tubulin (Abcam, ab15246; 1:1,000), anti-Vinculin (Sigma-Aldrich, V9131; 1:1,000), anti-ATG5 (Cell signaling, 8540; 1:1,000), anti-LC3 (Nano Tools, 0260-100/LC3-2G6; 1:100), anti-GABARAP (E1J4E, Cell Signaling, 13733; 1:1,000), anti-GABARAPL1 (Abcam, ab86497; 1:500), anti-P62/SQSTM1 (MBL, PM045; 1:1,000), anti-LAMP1 (Abcam, ab24170; 1:1,000), anti-LAMP2 (Abcam, ab13524; 1:500), anti-ATG4B (Cell signaling, 5299; 1:1,000), anti-ULK1 (Santa Cruz, sc-33182; 1:500), anti-Beclin-1 (Cell Signaling, 3738; 1:1,000) and anti-EGFR (D38B1, Cell Signaling, 4267; 1:1,000) antibodies were purchased and used according to the manufacturer’s recommendations.

Ebner P., Poetsch I., Deszcz L., Hoffmann T., Zuber J, & Ikeda F. (2018). The IAP family member BRUCE regulates autophagosome–lysosome fusion. Nature Communications, 9, 599.

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Autophagy Regulatory Protein Analysis

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The primary antibodies used in this study were as follows: the rabbit antibodies anti-LC3B, phospho-Ser79 ACC, total ACC, phospho-Thr172 AMPK, total AMPK, phospho-Thr389 p70S6K, total p70S6K and phospho-serin757 ULK1 (Cell Signaling); anti-ULK1 (Santa Cruz Biotechnology); anti-TPC1 and TPC2 (Abcam); anti-GAPDH (Calbiochem); anti-β-actin (Sigma-Aldrich).

Pereira G.J., Antonioli M., Hirata H., Ureshino R.P., Nascimento A.R., Bincoletto C., Vescovo T., Piacentini M., Fimia G.M, & Smaili S.S. (2016). Glutamate induces autophagy via the two-pore channels in neural cells. Oncotarget, 8(8), 12730-12740.

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