H ras

Manufactured by Santa Cruz Biotechnology

Sourced in United States

H-Ras is a small GTPase protein that plays a key role in cellular signal transduction pathways. It functions as a molecular switch, cycling between an active GTP-bound state and an inactive GDP-bound state. H-Ras is involved in the regulation of cell growth, differentiation, and survival.

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Lab products found in correlation

β actin, by Santa Cruz Biotechnology (4 mentions) Flag tag (flag), by Merck Group (3 mentions) β actin, by Merck Group (3 mentions) P erk, by Cell Signaling Technology (3 mentions) C myc, by Santa Cruz Biotechnology (2 mentions) α tubulin, by Merck Group (2 mentions) Pvdf membrane, by Thermo Fisher Scientific (2 mentions) Vinculin, by Merck Group (2 mentions) Prb 159p, by Merck Group (2 mentions) Horseradish peroxidase conjugated secondary antibody, by Santa Cruz Biotechnology (2 mentions) Ecl reagent, by Cytiva (2 mentions) Phosstop, by Roche (2 mentions) Idh1 r132h, by Dianova (2 mentions) β actin, by Cell Signaling Technology (2 mentions) Protease inhibitor cocktail, by Roche (2 mentions) P jnk, by Cell Signaling Technology (2 mentions) Stat3, by Thermo Fisher Scientific (1 mentions) P stat5, by Cell Signaling Technology (1 mentions) Stat5, by Cell Signaling Technology (1 mentions) Hsc70, by Santa Cruz Biotechnology (1 mentions)

Spelling variants of this product

Anti-H-Ras antibody (3 citations) Rabbit anti-H-RAS (2 citations) H‐Ras antibody (2 citations) Mouse anti-H-Ras (2 citations)

21 protocols using h ras

Western Blot Analysis of Protein Targets

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Western blots were performed as previously described [21] (link), using the following antibodies detecting: Akt (cat.# 4691, Cell signalling, dilution 1:1000), Dnmt1 (cat.# ab188453, Abcam; dilution 1:500), Dnmt3a (cat.# SC-20703, Santa Cruz; dilution 1:1000), Dnmt3b (cat.# PA1–884, Thermo Fisher; dilution 1:1000), Stat3 (cat.# SC-8019, Santa Cruz; dilution 1:500), P-Stat3 (cat.# 9145, Cell signalling, dilution 1:1000), Stat5 (cat.# 25656, Cell signalling, dilution 1:1000), P-Stat5 (cat.# 9359, Cell signalling, dilution 1:1000), mouse c-Met (cat.# SC-8057; Santa Cruz; dilution 1:500), c-Myc (cat.# SC-40, Santa Cruz; dilution 1:1000), Hsc-70 (cat.# SC-7298, Santa Cruz; dilution 1:10000), Lin28b (cat.# SC-293120, Santa Cruz; dilution 1:500), h-Ras (cat.# SC-520, Santa Cruz, dilution 1:1000) and β-actin (cat.# SC-130657, Santa Cruz; dilution 1:1000).

Lopusna K., Nowialis P., Opavska J., Abraham A., Riva A, & Opavsky R. (2021). Dnmt3b catalytic activity is critical for its tumour suppressor function in lymphomagenesis and is associated with c-Met oncogenic signalling. EBioMedicine, 63, 103191.

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In Vivo Ubiquitylation and Immunoblot Analysis

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In vivo ubiquitylation assays and immunoblot analyses were performed as previously described [19 (link)]. Briefly, N-ethylmaleimide (10 mM, Sigma) was added to the radioimmunoprecipitation assay (RIPA) buffer (Upstate Biotechnology). The lysates were incubated with the indicated antibodies and Protein G agarose at 4°C for 12 h, and the beads were washed three times with cold RIPA buffer. Immunoblot analysis was performed as previously described [19 (link)]. Primary antibodies were obtained from the following sources; anti-pan-Ras (Millipore); -K-Ras, -N-Ras, -H-Ras, -β-actin, -p-ERK, -p-AKT, -PKCδ, -PKCα,-PCNA, and -c-Myc (Santa Cruz Biotechnology); -HA, -PKCε, and -PKCζ (Cell Signaling Technology); -Flag (Sigma); and -ER-α36 (Cell Applications). Secondary antibodies were horseradish peroxidase (HRP)-conjugated anti-mouse (Cell Signaling Technology) and HRP-conjugated anti-rabbit (Bio-Rad).

Koo K.H., Jeong W.J., Cho Y.H., Park J.C., Min D.S, & Choi K.Y. (2015). K-Ras stabilization by estrogen via PKCδ is involved in endometrial tumorigenesis. Oncotarget, 6(25), 21328-21340.

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Western Blot Analysis of Cancer Signaling

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Western blot and protein detection was performed as previously described [43 (link)]. The following primary antibodies were used: RB, phospho-RB^S807/S811, CDC25C, phospho-AKT^S473, phospho-cyclin E1^T62, phospho-ERK1/2^T202/Y204, ERBB2, phospho-ERBB2^Y1221/1222 and MYC (Cell Signaling, Danvers, MA); BRCA1, p53, cyclin E, CDK1, CDK2, ETV4, ETV5 and HRAS (Santa Cruz Biotechnology, Santa Cruz, CA); Actin (Abcam, Cambridge, MA); GAPDH (Fitzgerald, Acton, MA); β-Actin (Sigma-Aldrich); PAX8 (Epitomics, Burlingame, CA);

Taylor-Harding B., Aspuria P.J., Agadjanian H., Cheon D.J., Mizuno T., Greenberg D., Allen J.R., Spurka L., Funari V., Spiteri E., Wang Q., Orsulic S., Walsh C., Karlan B.Y, & Wiedemeyer W.R. (2014). Cyclin E1 and RTK/RAS signaling drive CDK inhibitor resistance via activation of E2F and ETS. Oncotarget, 6(2), 696-714.

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Comprehensive Western Blot Analysis

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Whole six-well cells were lysed in 250ul 0.1% SDS and 10ul 1mM PMSF. Protein extracts were separated by SDS-PAGE and analyzed using the following primary antibodies: Id1 (Santa Cruz, sc-374287), Id3 (Santa Cruz, sc-56712), HRAS (Santa Cruz, sc-29), Vimentin (Santa Cruz, sc-6260), p-MEK (CST, 9154), p-ERK (CST, 4370), MEK (CST, 4694), ERK (CST, 4695), Snail (CST, 3879), Slug (CST, 9585), Slug (CST, 9585), MMP2 (CST, 4022), MMP9 (CST, 3852) , C-FOS (Proteintech, 66590-1-Ig), C-JUN (Proteintech, 24909-1-AP), LEF1(Abcam, ab137872), DUSP16 (Abcam, ab181088), E-cadherin(Abcam, ab40772), N-cadherin(Abcam, ab18203) and GADPH antibody (Abcam, ab8245). The next day, the membranes were incubated with secondary antibodies (CST,7076,7074) at room temperature for 2 hours. All results were repeated 3 times independently.

Wang X., Zhao Y., Fei X., Lu Q., Li Y., Yuan Y., Lu C., Li C, & Chen H. (2020). LEF1/Id3/HRAS axis promotes the tumorigenesis and progression of esophageal squamous cell carcinoma. International Journal of Biological Sciences, 16(13), 2392-2404.

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Antibody Characterization for DNA Damage Response

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The following antibodies were used for immunoblotting, immunofluorescence, or flow cytometry: SSRP1 (609702; Biolegend, San Diego, CA), SPT16 (607002; Biolegend), HRAS (sc-520; Santa Cruz, Dallas TX), p53 (ab27696, Abcam, Cambridge, United Kingdom), p21 (sc-6246; Santa Cruz), γH2AX (9718; Cell Signaling, Danvers, MA), RPA70 (2267; Cell Signaling), MCM7 (sc-9966; Santa Cruz), MCM4 (ab4459; Abcam), phospho-(Ser/Thr) ATM/ATR substrate antibody (2851; Cell Signaling), Chk1 (2360, Cell Signaling), p-Chk1-Ser317 (2344; Cell Signaling), p-Chk1-Ser296 (90178; Cell Signaling), cyclin A (4656; Cell Signaling), PCNA (2586; Cell Signaling), ssDNA (MAB3868; MilliporeSigma), Cross-Adsorbed Secondary Antibody, Alexa Fluor 488 (A-11008; Thermo Fisher Scientific, Waltham, MA), and β-actin (A3854, Sigma-Aldrich). Where indicated, cells were treated with 1 or 2.5 mM HU (H8627; Sigma-Aldrich). Cell cycle analysis was done using propidium iodide or Hoechst 33342 stain. CBL0137 (1 μM; Incuron, Buffalo, NY, USA) was used as a positive control for p53 activation and p21 induction.

Prendergast L., Hong E., Safina A., Poe D, & Gurova K. (2020). Histone chaperone FACT is essential to overcome replication stress in mammalian cells. Oncogene, 39(28), 5124-5137.

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Immunoblotting Antibody Detection Protocol

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Immunoblotting was performed as described previously [31 (link)]. The following antibodies were used: H-Ras (Santa Cruz), SV40 Large-T (Santa Cruz), FLAG-Tag (Sigma), hTERT (Epitomics), Ku 70 (Cell Signaling), Ku 80 (Cell Signaling), α-tubulin (Sigma) and β-actin (Abcam).

Cao X., Kong C.M., Mathi K.M., Lim Y.P., Cacheux-Rataboul V, & Wang X. (2014). The use of transformed IMR90 cell model to identify the potential extra-telomeric effects of hTERT in cell migration and DNA damage response. BMC Biochemistry, 15, 17.

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Protein Analysis by Immunoblotting

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Cell pellets were lysed and sonicated in EBC lysis buffer (50 mM Tris-Cl, pH7.4, 120mM NaCl, 0.5% NP-40, 1mM EDTA) containing HALT Protease and Phosphatase Inhibitor cocktail (Thermo Scientific) and 1mM phenylmethylsulfonyl fluoride (PMSF). Thirty μg of protein were separated on SDS-polyacrylamide gels. Proteins were transferred to PVDF (Millipore) and probed with antibodies. ARF (mouse), Actin, p53 (human), Gamma-tubulin, H-Ras, ISG15 (human), and STAT1 were all purchased from Santa Cruz Biotechnologies. p53 (mouse), phospho-STAT1^Tyr701, phospho-STAT1^Ser727, phospho-STAT3^Tyr705, and STAT3 were purchased from Cell Signaling. ARF (human) and GAPDH were purchased from Bethyl Laboratories, and MDM2 was from Millipore. The mouse ISG15 antibody was a gift from Dr. Deborah Lenschow. Secondary horseradish peroxidase conjugated antibodies (Jackson Immunoresearch) were used and ECL plus was used to visualize the bands (GE Healthcare).

Forys J.T., Kuzmicki C.E., Saporita A.J., Winkeler C.L., Maggi LB J.r, & Weber J.D. (2014). ARF and p53 coordinate tumor suppression of an oncogenic IFN-β-STAT1-ISG15 signaling axis. Cell reports, 7(2), 514-526.

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Western Blot Analysis of Protein Expression

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Lysates of cells and tissues were prepared in SDS sample buffer (60 mM Tris-HCl at pH 6.8, 10% glycerol, 2% SDS, and 5% 2-mercaptoethanol). Equal amounts proteins (25 μg) were separated on 4% to 12% SDS-PAGE gels, transferred to PVDF membranes (Invitrogen), and incubated with primary antibodies specific to ERK (Cell Signaling; 4695), P-ERK (Cell Signaling; 4370S), H-RAS (Santa Cruz; sc-520), Krt10 (Covance; PRB-159P), Vinculin (Sigma; V4139). Blots were rinsed in TBST and incubated in peroxidase-conjugated anti-mouse or anti-rabbit secondary antibodies, respectively. Proteins were visualized using an ECL detection system (Pierce Biotech) and exposed to film. All experiments were repeated in triplicate.

Palazzo E., Kellett M., Cataisson C., Gormley A., Bible P.W., Pietroni V., Radoja N., Hwang J., Blumenberg M., Yuspa S.H, & Morasso M. (2015). The homeoprotein DLX3 and tumor suppressor p53 co-regulate cell cycle progression and squamous tumor growth. Oncogene, 35(24), 3114-3124.

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Western Blot Analysis of Key Cellular Proteins

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Cells were washed in cold PBS and lysed in NP40 cell lysis buffer (Life Technologies) supplemented with 1× PhosStop and protease inhibitor cocktail (Roche). The protein was quantitated and used (30 μg) for Western blot analysis using primary antibodies against IDH1 (Dianova), IDH1R132H (Dianova), H-Ras (Santa Cruz Biotechnology), RAD51 (EMD Chemicals, PC130), MGMT, or β-actin (Cell Signaling Technologies) and the appropriate horseradish peroxidase–conjugated secondary antibodies (Santa Cruz Biotechnology). Antibody binding was detected using ECL reagents (Amersham Pharmacia Biotech).

Ohba S., Mukherjee J., See W.L, & Pieper R.O. (2014). Mutant IDH1-driven cellular transformation increases RAD51-mediated homologous recombination and temozolomide resistance. Cancer research, 74(17), 4836-4844.

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Comprehensive Western Blot Analysis

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Cells were lysed in RIPA lysis buffer (Life Technologies) supplemented with 1x PhosStop and protease inhibitor cocktail (Roche). Protein (30 μg) was used for Western blot analysis using primary antibodies against IDH1 (Dianova), IDH1R132H (Dianova), H-Ras (F235, Santa Cruz Biotechnology), p53 (Cell Signaling), HPV16E7 (Santa Cruz Biotechnology), ATRX (Cell Signaling), β-actin (Cell Signaling), H3K4me3, H3K9me2, H3K9me3, H3K27me3 (all Active Motif), or Histone H3 (Abcam) and the appropriate horseradish peroxidase-conjugated secondary antibodies (Santa Cruz Biotech). Antibody binding was detected using ECL reagents (Amersham).

Ohba S., Mukherjee J., Johannessen T.C., Mancini A., Chow T.T., Wood M., Jones L., Mazor T., Marshall R.E., Viswanath P., Walsh K.M., Perry A., Bell R.J., Phillips J.J., Costello J.F., Ronen S.M, & Pieper R.O. (2016). Mutant IDH1 expression drives TERT promoter reactivation as part of the cellular transformation process. Cancer research, 76(22), 6680-6689.

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