Anti p chk1

Manufactured by Cell Signaling Technology

Sourced in United States, Canada

Anti-p-CHK1 is a laboratory product offered by Cell Signaling Technology. It is a primary antibody that detects phosphorylated Checkpoint Kinase 1 (CHK1) protein. CHK1 is a key regulator of the cell cycle and DNA damage response.

Automatically generated - may contain errors

Lab products found in correlation

Spelling variants of this product

P-Chk1 (77 citations) Anti-CHK1 (55 citations) Anti-P-Chk1-Ser317 (6 citations)

24 protocols using anti p chk1

Immunoblotting of Keratinocyte Signaling Pathways

Check if the same lab product or an alternative is used in the 5 most similar protocols

The inhibitors used in this study were as follows: 100 nM UCN01 and 5 µM VE822 (Selleckchem, Houston, TX); 10 µM pimozide (Sigma-Aldrich, St. Louis, MO); and 5 µM CHK2i (Calbiochem, San Diego, CA). The antibodies used in this study were as follows: antiinvolucrin, anti-TopBP1, and anti-GAPDH (Santa Cruz, Santa Cruz, CA) and anti-STAT-5, anti-CHK2, anti-ATM, anti-p-CHK2 (Thr68), anti-p-ATM (Ser1981), anti-CHK1, anti-ATR, anti-p-CHK1 (Ser296), anti-p-CHK1 (Ser345), and anti-p-ATR (Cell Signaling, Inc., Danvers, MA). For Western blot analysis, cell lysates were processed as previously described (36 (link)). Briefly, keratinocytes were first isolated from J2 feeders by treatment with Versene (phosphate-buffered saline [PBS] containing 0.5 mM EDTA) and lysed in radioimmunoprecipitation assay (RIPA) lysis buffer on ice for 30 min. The protein samples were separated on SDS-PAGE gels and then transferred to polyvinylidene difluoride (PVDF) membranes. The membranes were finally developed using Enhanced Chemiluminescence (ECL) Prime or ECL reagents (Amersham, Pittsburgh, PA). Chemiluminescence signals were detected using Eastman Kodak X-ray films.

Hong S., Cheng S., Iovane A, & Laimins L.A. (2015). STAT-5 Regulates Transcription of the Topoisomerase IIβ-Binding Protein 1 (TopBP1) Gene To Activate the ATR Pathway and Promote Human Papillomavirus Replication. mBio, 6(6), e02006-15.

+ Open protocol

+ Expand

Immunoblotting Analysis of DNA Damage Signaling

Check if the same lab product or an alternative is used in the 5 most similar protocols

Lysates were subjected to immunoblotting with anti-MUC1-C (HM-1630-P1ABX; Thermo Fisher Scientific), anti-β-actin (A5441; Sigma), anti-lamin A/C (4777), anti-γH2AX (9718), anti-ATR (2790), anti-pCHK1 (2348), anti-CHK1 (2360), anti-pBRCA1 (9009), anti-BRCA1 (14823), anti-BMI1 (6964), anti-H2AUb1 (8240), anti-H2A (12349), anti-EZH2 (5246), anti-H3K27me3 (9733), anti-H3 (9715), anti-H3K56ac (4243), anti-PARP1 (9532) (Cell Signaling Technology, Danvers, MA, USA), anti-pATR (GTX128145; GeneTex, Irvine, CA, USA) and anti-FANCD2 (ac108928; Abcam, Cambridge, CB4, 0Fl, UK). Signal intensity was determined using ImageJ 1.51k software (NIH, Bethesda, MD, USA).

Yamamoto M., Jin C., Hata T., Yasumizu Y., Zhang Y., Hong D., Maeda T., Miyo M., Hiraki M., Suzuki Y., Hinohara K., Rajabi H, & Kufe D. (2019). MUC1-C INTEGRATES CHROMATIN REMODELING AND PARP1 ACTIVITY IN THE DNA DAMAGE RESPONSE OF TRIPLE-NEGATIVE BREAST CANCER CELLS. Cancer research, 79(8), 2031-2041.

+ Open protocol

+ Expand

Western Blot Analysis of DNA Damage Signaling

Check if the same lab product or an alternative is used in the 5 most similar protocols

Cells and tumor fragments were lysed in a buffer (Mesoscale discovery, #R60TX-2) with protease and phosphatase inhibitors (Thermo Fisher Scientific, #78437; Thermo Fisher Scientific, #78420). Lysates were separated by SDS-PAGE (Bio-Rad, #5678085); then transferred to polyvinylidene fluoride membranes. Membranes were incubated with primary antibodies: anti-pCHK1 (Ser345; Cell Signaling Technology, #2348), anti-pKAP1(S824; Abcam; catalog no. ab133440), anti-KAP1 (Abcam; catalog no. 10484), anti-ATM (Novus; NB100–104SS) and anti-pDNA-PKcs (Abcam; catalog no. 124918), anti-CHK1 (Cell Signaling Technology, #2348), or anti-γH2AX (EMD Millipore; catalog no. 05–636). Signal detection was carried out with secondary anti-rabbit (Jackson Immuno Research, #711–035–152) or anti-mouse (Jackson Immuno Research, #115–035–174) horseradish peroxidase (HRP) antibody and SuperSignal West Dura Extended Duration Substrate (Thermo Fisher Scientific, #34076) using a Bio-Rad ChemiDoc Touch Imaging System (Bio-Rad, #1708370).

Roulston A., Zimmermann M., Papp R., Skeldon A., Pellerin C., Dumas-Bérube É., Dumais V., Dorich S., Fader L.D., Fournier S., Li L., Leclaire M.E., Yin S.Y., Chefson A., Alam H., Yang W., Fugère-Desjardins C., Vignini-Hammond S., Skorey K., Mulani A., Rimkunas V., Veloso A., Hamel M., Stocco R., Mamane Y., Li Z., Young J.T., Zinda M, & Black W.C. (2021). RP-3500: A Novel, Potent, and Selective ATR Inhibitor that is Effective in Preclinical Models as a Monotherapy and in Combination with PARP Inhibitors. Molecular Cancer Therapeutics, 21(2), 245-256.

+ Open protocol

+ Expand

Comprehensive Antibody Catalog for Research

Check if the same lab product or an alternative is used in the 5 most similar protocols

In this work, the following antibodies were used: primary antibodies: anti-hnRNP UL1 (Abcam #ab68480 or Santa Cruz Biotechnology #sc-393434), anti-FLAG (Sigma #A8592), anti-Actin (MP #691001), anti-Fibrillarin (Santa Cruz Biotechnology #sc-25397), anti-Nucleolin (Abcam #ab22758), anti-FUS (Santa Cruz Biotechnology #sc-47711), anti-γH2A.X (Santa Cruz Biotechnology #sc-517348), anti-RPS6 (Abcam #ab70227), anti-RPS15 (Antikoerper #ABIN2786563), anti-RPA32 (Bethyl Laboratories #A300-245A), anti-pChk1 (Cell Signaling Technology #2341), anti-XRCC1 (Invitrogen #MA5-13412), anti-53BP1 (Abcam #ab175933), anti-Rad50 (Abcam #ab124682), anti-RPA194 (Santa Cruz Biotechnology #sc-46699), normal mouse IgG (Santa Cruz Biotechnology #sc-2025), and anti-digoxygenin-AP Fab fragments (Roche #11093274910); secondary antibodies: goat anti-mouse IgG-horseradish peroxidase (HRP) (Santa Cruz Biotechnology #sc-516102), goat anti-rabbit IgG-HRP (Santa Cruz Biotechnology #sc-2004), anti-mouse Alexa Fluor 555 (Thermo Fisher Scientific #A21422), anti-mouse Alexa Fluor 488 (Thermo Fisher Scientific #A32723), anti-rabbit Alexa Fluor 555 (Thermo Fisher Scientific #A32732) or anti-rabbit Alexa Fluor 488 (Thermo Fisher Scientific #A32731).

Cichocka M., Karlik A., Plewka P., Gawade K., Stępień A., Świergiel P., Gadgil A, & Raczyńska K.D. (2022). The Novel Role of hnRNP UL1 in Human Cell Nucleoli. International Journal of Biological Sciences, 18(13), 4809-4823.

+ Open protocol

+ Expand

Western Blotting of Signaling Proteins

Check if the same lab product or an alternative is used in the 5 most similar protocols

Proteins were extracted from SK-N-MC cells using SDS sample buffer and boiled at 99°C for 10 min prior to western blotting. Primary antibodies used were anti-NUDT16 (SAB2107004, 1:2,000) from Sigma-Aldrich; anti-p-ATR (2853, 1:1,000), anti-p-ATM (5883, 1:1,000), anti-p-CHK1 (2348, 1:1,000), anti-p-CHK2 (2197, 1:1,000), anti-cleaved caspase 3 (9664, 1:500), and anti-myc (2276, 1:2,000) from Cell Signaling Technology; anti-Dicer (ab14601, 1:2,000), anti-TDP-43 (ab109535, 1:2,000), and anti-β-tubulin (ab6046, 1:2,000) from Abcam; and anti-GFP (JL-8, 1:4,000) from Clontech. Secondary antibodies were goat anti-rabbit (11-035-045, 1:5,000) and goat anti-mouse (115-035-062, 1:5,000) from Jackson ImmunoResearch.

Peng S.I., Leong L.I., Sun J.K., Chen Z.S., Chow H.M, & Chan H.Y. (2022). A peptide inhibitor that rescues polyglutamine-induced synaptic defects and cell death through suppressing RNA and protein toxicities. Molecular Therapy. Nucleic Acids, 29, 102-115.

+ Open protocol

+ Expand

Antibodies for DNA Damage Response Analysis

Check if the same lab product or an alternative is used in the 5 most similar protocols

The following antibodies were used: anti-TIMELESS (Bethyl Laboratories, A300-961A and A300-960A, the latter used for IP), anti-PARP1 (Invitrogen, 436400), anti-PARP1 (Cell Signaling Technology, 9542 and 46D11, the latter used for IF), anti-FLAG M2 (Sigma, F3165), anti-p-Chk1 (Cell Signaling, 2344), anti-p-Chk2 (Cell Signaling, 2661), anti-p-ATM (Cell Signaling, 4526), anti-LIG3 (Bethyl, A301-637A), anti-HLTF (Bethyl, A300-229A), anti-KU70 (Santa Cruz Biotechnology, sc-9033), anti-KU80 (Neomarkers, MS-285-P1; Cell Signaling, 2180), anti-XRCC1 (Cell Signaling, 2735), anti-DNA-PK (Santa Cruz, sc-5282), anti-DNA-PK (Cell Signaling, 12311), anti-TIPIN (Bethyl, A301-474A-1), anti-RPA1 (Santa Cruz, sc46504), anti-RPA2 (Millipore, 04-1481), anti-SSRP1 (Abcam, ab26212), anti-SPT16 (Cell Signaling, 12191), anti-SKP1 (Santa Cruz, sc-5281), and anti-phospho-H2AX (Millipore, 05-636).

Young L.M., Marzio A., Perez-Duran P., Reid D.A., Meredith D.N., Roberti D., Star A., Rothenberg E., Ueberheide B, & Pagano M. (2015). TIMELESS Forms a Complex with PARP1 Distinct from Its Complex with TIPIN and Plays a Role in the DNA Damage Response. Cell reports, 13(3), 451-459.

+ Open protocol

+ Expand

H2O2-Induced Stress Response Signaling

Check if the same lab product or an alternative is used in the 5 most similar protocols

Forty‐eight hours after transfection, cells were induced by 500 μM H₂O₂ for 24 h. Then, all cells were homogenized in RIPA lysis buffer (Promega) on ice for 30 min. Protein concentrations were estimated using the BCA protein assay (Thermo). Equal amounts of cell lysates were loaded and separated on 12% SDS‐PAGE gels, and proteins were electrotransferred to nitrocellulose membranes, as previously described. The membranes were incubated with a 1:1000 dilution of anti‐hnRNPK, anti‐p53, anti‐p21, anti‐γH2A.X, anti‐pCHK1, anti‐pCHK2, anti‐cyclin D1(Cell Signal Technology), and anti‐β‐actin (1:10000 dilution, Santa Cruz, CA) antibodies overnight at 4°C in 5% nonfat milk, followed by washing three times with TBS‐T buffer and incubation with horseradish peroxidase‐conjugated goat anti‐rabbit IgG (1:10000 dilution with 5% nonfat milk) for 45 min at room temperature. After washing three times with TBS‐T buffer, membranes were visualized using the ECL detection system (Thermo Fisher Scientific).

Li M., Yang X., Zhang G., Wang L., Zhu Z., Zhang W., Huang H, & Gao R. (2022). Heterogeneous nuclear ribonucleoprotein K promotes the progression of lung cancer by inhibiting the p53‐dependent signaling pathway. Thoracic Cancer, 13(9), 1311-1321.

+ Open protocol

+ Expand

Western Blot Analysis of Apoptosis Markers

Check if the same lab product or an alternative is used in the 5 most similar protocols

The protein samples were prepared as described previously. Briefly, proteins of HL60 cells were extracted in lysis buffer (150.0 mM NaCl, 50 mM Tris–HCl, 1 mM EDTA, 0.1 % SDS, 0.5 % dexoycholic acid, 0.02 % sodium azide, 1 % NP-40, 2.0 μg/ml aprotinin, 1 mM phenylmethylsulfonylfluoride). The lysates were centrifuged at 10⁴ × g for 15 min at 4 °C. Equivalent amounts of proteins were analyzed by 8 %-15 % SDS-PAGE and electroblotted onto PVDF membranes (Millipore Corporation, Billerica, Massachusetts), and probed with primary antibodies. Appropriate antibodies to anti-caspase-3, anti-poly-ADP-ribose polymerase (PARP), anti-Bax, anti-chk1, anti-chk2 and anti-β-actin from Santa Cruz Biotechnology (Santa Cruz, CA, USA); anti-Bcl-2, anti-p-chk1, anti-p-chk2, and anti-γ-H2AX from Cell Signaling Technology (Beverly, MA, USA); and anti-Cytochrome C from Cell Signaling Technology (Boston, MA, USA) were used. The proteins were visualized with peroxidase-coupled secondary antibodies (Southern Biotech, Birmingham, UK), and using the enhanced chemiluminescence detection system (Biological Industries, Beit Haemek, Israel) for detection.

Xu Z., Shao J., Li L., Peng X., Chen M., Li G., Yan H., Yang B., Luo P, & He Q. (2016). All-trans retinoic acid synergizes with topotecan to suppress AML cells via promoting RARα-mediated DNA damage. BMC Cancer, 16, 2.

+ Open protocol

+ Expand

High-Throughput Screening of DNA Damage Response

Check if the same lab product or an alternative is used in the 5 most similar protocols

The compound library was obtained from the National Cancer Institute and was composed of 933 compounds including natural product compounds and bioactive compounds (Natural Compounds Set III and 1 mM Mechanistic Diversity Set II). Monoclonal antibodies for western blotting against phosphorylated RPA (p-RPA),RPA,and CTIP were purchased from Bethyl (Montgomery,TX); anti-p-CHK1 and anti-CHK1 antibodies were purchased from Cell Signaling Technology, Inc. (Beverly, MA); anti-BRCA1 antibody was purchased from Santa Cruz Biotechnology, Inc. (Dallas,TX);and anti-tubulin antibody was purchased from Sigma (St. Louis, MO). Goat anti-mouse IgG-HRP and goat anti-rabbit IgG-HRP were purchased from Santa Cruz Biotechnology, Inc. Primary antibody for immunofluorescent staining against Rad51 (SC-8349) and 53BP1 (SC-22760) were purchased from Santa Cruz Biotechnology, Inc., anti-p-RPA32 (A300-245A)was purchased from Bethyl, anti-γ-H2AX (16-202A) was purchased from EMD Millipore. Secondary antibody Alex Fluor-conjugated goat anti-rabbit IgG (A21206) and secondary antibody Alex Fluor-conjugated goat anti-mouse IgG (A-11001) were purchased from Life Technology (Waltham, MA).

Zhang L., Peng Y., Uray I.P., Shen J., Wang L., Peng X., Brown P.H., Tu W, & Peng G. (2017). Natural product β-thujaplicin inhibits homologous recombination repair and sensitizes cancer cells to radiation therapy. DNA repair, 60, 89-101.

+ Open protocol

+ Expand

Western Blot Analysis of DNA Damage Response Proteins

Check if the same lab product or an alternative is used in the 5 most similar protocols

The antibodies from this study were as follows: anti-keratin-10, anti-loricrin, anti-TopBP1, and anti-GAPDH (Santa Cruz, Santa Cruz, CA); anti-E2F1, anti-p63, anti-ATR, anti-p-ATR, anti-CHK1, anti-p-CHK1, anti-CHK2, anti-p-CHK2, anti-RAD51, anti-Mre11, anti-NBS1, anti-BRCA1, and anti-BRCA2 (Cell Signaling, Danvers, MA); anti-p73 (Abcam, Cambridge, MA); anti-p53 (Millipore, Burlington, MA); anti-p-TopBP1 (Abgent, San Diego, CA). AKT inhibitors MK2206 (2μM) and LY294002 (25μM) were purchased from SelleckChem (Houston, TX). Cell lysates were processed and assayed by western blot analysis as previously described ^{60 (link)}. Briefly, J2 feeders were firstly removed by Versene (PBS containing 0.5 mM EDTA) treatment and keratinocytes were collected and lysed in RIPA lysis buffer on ice for 30 minutes. The samples were then electrophoresed on SDS-page gels and transferred to PVDF membranes. At last the membranes were developed using ECL prime or ECL reagents (Amersham, Pittsburgh, PA) and chemiluminescence signals were detected using Eastman Kodak x-ray films.

Hong S., Xu J., Li Y., Andrade J., Hoover P., Kaminski P.J, & Laimins L.A. (2019). Topoisomerase IIβ-binding protein 1 activates expression of E2F1 and p73 in HPV positive cells for genome amplification upon epithelial differentiation. Oncogene, 38(17), 3274-3287.

+ Open protocol

+ Expand

About PubCompare

Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.

We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.

However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.

Ready to get started?

Revolutionizing how scientists
search and build protocols!