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H 125

Manufactured by Santa Cruz Biotechnology
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The H-125 is a high-quality laboratory equipment designed for performing basic centrifugation tasks. It features a compact and durable construction, allowing for efficient and reliable operation within the laboratory environment.

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Lab products found in correlation

Ab13822, by BD (2 mentions) Alexa fluor 488, by Thermo Fisher Scientific (2 mentions) Hpa018425, by Merck Group (2 mentions) F3648, by Merck Group (2 mentions) Mab1378, by Merck Group (2 mentions) Nki goh3, by Merck Group (2 mentions) Twist2c1a, by Merck Group (2 mentions) T6451, by Merck Group (2 mentions) Anti mouse, by Jackson ImmunoResearch (2 mentions) Anti rabbit, by Jackson ImmunoResearch (2 mentions) Anti chicken, by Jackson ImmunoResearch (2 mentions) Ab50887, by Merck Group (2 mentions) Anti e cadherin, by Abcam (2 mentions) Anti e cadherin, by BD (2 mentions) Ab103580, by Abcam (2 mentions) Ab11042, by Abcam (2 mentions) Ep436y, by Abcam (2 mentions) Ecl plus kit, by Cytiva (2 mentions) Anti tap63, by BioLegend (2 mentions) Img 246, by Novus Biologicals (2 mentions)

10 protocols using h 125

1

Immunofluorescence and Immunoblot Analysis of YAP/TAZ

Cited 1 time
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For immunofluorescence, 1/200 rabbit anti-YAP antibody (H-125, Santa Cruz), 1/400 rabbit anti-YAP/TAZ antibody (D24E4, Cell Signaling), 1/200 phalloidin-AlexaFluor 594 (Life Technologies), and 1/1000 anti-rabbit IgG-AlexaFluor 488 conjugate (Life Technologies) were used. For immunoblot, 1/3000 rabbit anti-YAP antibody (H-125, Santa Cruz), 1/3000 rabbit anti-TAZ antibody (#2149, Cell Signaling), 1/3000 rabbit anti-YAP/TAZ antibody (D24E4, Cell Signaling), 1/10,000 mouse anti-GAPDH antibody (6C5, Millipore), 1/5000 anti-mouse IgG-HRP (GE Healthcare), and 1/5000 anti-rabbit IgG-HRP (GE Healthcare), were used. Antibodies for Western blot were diluted in Can Get Signal reagents (Toyobo). Western blot using standard SDS–PAGE gel or gels containing Phos-tag-acrylamide (SuperSep Phos-tag, Wako) was performed as previously described [18] (link).
Oku Y., Nishiya N., Shito T., Yamamoto R., Yamamoto Y., Oyama C, & Uehara Y. (2015). Small molecules inhibiting the nuclear localization of YAP/TAZ for chemotherapeutics and chemosensitizers against breast cancers. FEBS Open Bio, 5, 542-549.
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2

Antibody Panel for EMT Markers

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Primary antibodies include anti-β-actin (Abcam, ab13822, 1:3000), anti-E-cadherin (BD, 610182, 1:200 for immunostaining, 1:1000 for Western blotting), anti-E-cadherin (Abcam, ab11512, Decma-1, 1:200), anti-G3BP2 (Sigma-Aldrich , HPA018425, 1:200, 1:1000), anti-Fibronectin (Sigma-Aldrich, F3648, 1:200), anti-Integrin α6 (Millipore, MAB1378, NKI-GoH3, 1:200), anti-human Laminin V (Chemicon, D4B5, 1:200), anti-mouse Laminin V (kind gift from M. Aumailley, 1:1000), anti-Twist1 (Santa Cruz, ab50887, Twist2C1a, 1:100, 1:1000), Rabbit anti-Twist1 (Sigma-Aldrich, T6451, 1:1000), 5b7 mouse anti-Twist1 hybridoma cell line (1:1000), anti-YAP1 (Santa Cruz, H-125, 1:100). AIIB2 hybridoma supernatant was used for β1 integrin blocking experiments (Developmental Studies Hybridoma Bank, 1:1000). Secondary fluorescent antibodies used include anti-mouse, anti-rat, and anti-rabbit conjugated with Alexafluor 488, 546, and 647 (Life Technologies). Secondary horseradish peroxidase (HRP) conjugated antibodies used include anti-mouse, anti-rabbit, and anti-chicken (Jackson Immunoresearch).
Wei S.C., Fattet L., Tsai J.H., Guo Y., Pai V.H., Majeski H.E., Chen A.C., Sah R.L., Taylor S.S., Engler A.J, & Yang J. (2015). Matrix stiffness drives Epithelial-Mesenchymal Transition and tumour metastasis through a TWIST1-G3BP2 mechanotransduction pathway. Nature cell biology, 17(5), 678-688.
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3

Western Blot Analysis of Cell Signaling

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Fifty micrograms of protein were electrophoresed on a 10% or 15% SDS PAGE and transferred to PVDF membrane as described previously 4 (link),8 (link),34 (link). Blots were probed with anti-p63 (1:500) (4A4, Santa Cruz), anti-TAp63 (1:1000) (BioLegend), anti-TAp73 (1:500)(IMG-246, Imgenex), anti-p73 (Mouse) (1:250)(IMG-259A, Imgenex), anti-p73 (1:1000) (human) (EP436Y, Abcam), anti-p53 (WT) (1:1000)(CM5, Vector Labs), anti-IAPP (1:1000)(ab103580, Abcam), anti-His (1:1000)(G18, Santa Cruz), anti-Hexokinase II (1:10000)(C64G5, Cell Signaling), anti-calcitonin receptor (1:1000)(ab11042, Abcam), RAMP3(1:1000)(H125, Santa Cruz), and cleaved caspase 3 (1:1000)(Asp 175, Cell Signaling), at 4°C for 18 hours followed by incubation for one hour at room temperature with the appropriate secondary antibodies conjugated to horseradish peroxidase (1:5000)(Jackson Lab). β-actin (Sigma 1:5000) was used as a loading control. Detection was performed using the ECL Plus Kit (Amersham) following the manufacturer’s protocol and x-ray autoradiography.
Venkatanarayan A., Raulji P., Norton W., Chakravarti D., Coarfa C., Su X., Sandur S.K., Ramirez M.S., Lee J., Kingsley C.V., Sananikone E.F., Rajapakshe K., Naff K., Parker-Thornburg J., Bankson J.A., Tsai K.Y., Gunaratne P.H, & Flores E.R. (2014). IAPP driven metabolic reprogramming induces regression of p53 - deficient tumours in vivo. Nature, 517(7536), 626-630.
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4

Rat Brain Immunohistochemistry Protocol

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Following behavioral testing, rats were deeply anesthetized and transcardially perfused. Brain tissue was sectioned (30 μm) on a cryostat as previously detailed (Bangasser et al., 2013a (link)). Slices from the dose-response curve study were processed with cresyl violet to confirm placement. Sections from the rats in the subsequent studies were processed for immunohistochemistry as previously described (Bangasser et al., 2013b (link)). Briefly, every 4th section throughout the brain was quenched (0.75% H2O2, 20 min), blocked (phosphate buffered saline, triton, and 0.4% bovine serum albumin, 30 min), and then incubated for 48 hours with anti-cFOS (1:1000, Santa Cruz H-125). Following rinses, sections were incubated (90 min) in donkey anti-rabbit conjugated to biotin (1:200, Jackson ImmunoResearch). Then sections were rinsed and incubated (90 min) in avidin-biotin complex reagent using a kit (ABC Vectastain, Burlingame, CA). Sections were then rinsed and processed for diaminobenzadine (DAB; Vector Laboratories, Inc. SK-4100). Finally, sections were then rinsed, dehydrated, mounted, and cover-slipped with Permount.
Wiersielis K.R., Wicks B., Simko H., Cohen S.R., Khantsis S., Baksh N., Waxler D.E, & Bangasser D.A. (2016). Sex differences in corticotropin releasing factor-evoked behavior and activated networks. Psychoneuroendocrinology, 73, 204-216.
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5

Depletion of Abundant Proteins in Biological Samples

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FBS, pPEDF, ascites and “ascites-PEDF” were loaded on a Pierce Top 2 abundant proteins depletion column (ThermoFisher Scientific, Switzerland) following the provider’s recommendation to remove albumin and immunoglobulins before Western blot analysis. Proteins (10 μl FBS or ascites equivalent) were fractionated by SDS-PAGE and transferred to nitrocellulose membrane for immunoblot analysis using rabbit anti-PEDF antibodies (H-125, 1:1000 dilution from Santa Cruz Biotechnology, Germany) as primary antibodies and goat anti-rabbit IgG (H+L)-HRP conjugate (170-6515, 1:3000 dilution from Bio-Rad, Basel, Switzerland) as secondary antibodies. Specific signal was detected using Amersham ECL Prime Western Blotting Detection Reagent (Ge Healthcare, UK).
Ribaux P., Britan A., Thumann G., Delie F., Petignat P, & Cohen M. (2019). Malignant ascites: a source of therapeutic protein against ovarian cancer?. Oncotarget, 10(57), 5894-5905.
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6

Chromatin Immunoprecipitation (ChIP) Assay

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DNA in cells from two confluent 100-mm culture dishes (∼2 × 107 cells total) was pretreated with 1.5 mM ethylene glycol bis(succinimidylsuccinate) (Sigma) for 30 min at room temperature to capture proteins indirectly bound to DNA, and then crosslinked by incubating with 1% formaldehyde for 15 min. After DNA crosslinking, cells were sonicated by Bioruptor (BMS Co.) in SDS lysis buffer (50 mM Tris-Cl pH 8.0, 1% SDS and 10 mM EDTA) and diluted 10-fold with dilution buffer (16.7 mM Tris-Cl pH 8.0, 167 mM NaCl, 1.1% Triton X-100 and 1.2 mM EDTA) and processed for ChIP assays using 2 μg of anti-YAP antibody (H-125, Santa Cruz Biotechnology), anti-TEAD4 antibody (Abcam) or anti-SRF antibody (Cell Signaling) and Protein A/G agarose (GenDEPOT). YAP 5SA is used for maximal efficiency in YAP binding to the chromatin. See Supplementary Table 1 for primers used.
Kim T., Yang S.J., Hwang D., Song J., Kim M., Kyum Kim S., Kang K., Ahn J., Lee D., Kim M.Y., Kim S., Seung Koo J., Seok Koh S., Kim S.Y, & Lim D.S. (2015). A basal-like breast cancer-specific role for SRF–IL6 in YAP-induced cancer stemness. Nature Communications, 6, 10186.
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7

Western Blot Analysis of Cell Signaling

Cited 3 times
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Fifty micrograms of protein were electrophoresed on a 10% or 15% SDS PAGE and transferred to PVDF membrane as described previously 4 (link),8 (link),34 (link). Blots were probed with anti-p63 (1:500) (4A4, Santa Cruz), anti-TAp63 (1:1000) (BioLegend), anti-TAp73 (1:500)(IMG-246, Imgenex), anti-p73 (Mouse) (1:250)(IMG-259A, Imgenex), anti-p73 (1:1000) (human) (EP436Y, Abcam), anti-p53 (WT) (1:1000)(CM5, Vector Labs), anti-IAPP (1:1000)(ab103580, Abcam), anti-His (1:1000)(G18, Santa Cruz), anti-Hexokinase II (1:10000)(C64G5, Cell Signaling), anti-calcitonin receptor (1:1000)(ab11042, Abcam), RAMP3(1:1000)(H125, Santa Cruz), and cleaved caspase 3 (1:1000)(Asp 175, Cell Signaling), at 4°C for 18 hours followed by incubation for one hour at room temperature with the appropriate secondary antibodies conjugated to horseradish peroxidase (1:5000)(Jackson Lab). β-actin (Sigma 1:5000) was used as a loading control. Detection was performed using the ECL Plus Kit (Amersham) following the manufacturer’s protocol and x-ray autoradiography.
Venkatanarayan A., Raulji P., Norton W., Chakravarti D., Coarfa C., Su X., Sandur S.K., Ramirez M.S., Lee J., Kingsley C.V., Sananikone E.F., Rajapakshe K., Naff K., Parker-Thornburg J., Bankson J.A., Tsai K.Y., Gunaratne P.H, & Flores E.R. (2014). IAPP driven metabolic reprogramming induces regression of p53 - deficient tumours in vivo. Nature, 517(7536), 626-630.
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8

Molecular Profiling of Cancer Specimens

Cited 1 time
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All specimens were acquired from individuals with NSCLC and melanoma under the auspices of institutional review board (IRB)-approved clinical protocols at each hospital in which informed consent was obtained. BRAF and KRAS mutation status was assessed by established clinical DNA sequencing assays. Immunohistochemistry for YAP (H-125, Santa Cruz Biotechnology) was conducted on formalin-fixed, paraffin-embedded tumor sections as previously described33 (link),34 (link). Statistical significance was assessed and is reported as the P value from the χ2 test, with P < 0.05 considered significant.
Lin L., Sabnis A.J., Chan E., Olivas V., Cade L., Pazarentzos E., Asthana S., Neel D., Yan J.J., Lu X., Pham L., Wang M.M., Karachaliou N., Cao M.G., Manzano J.L., Ramirez J.L., Torres J.M., Buttitta F., Rudin C.M., Collisson E.A., Algazi A., Robinson E., Osman I., Muñoz-Couselo E., Cortes J., Frederick D.T., Cooper Z.A., McMahon M., Marchetti A., Rosell R., Flaherty K.T., Wargo J.A, & Bivona T.G. (2015). The Hippo effector YAP promotes resistance to RAF- and MEK-targeted cancer therapies. Nature genetics, 47(3), 250-256.
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9

Antibody Panel for EMT Markers

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Primary antibodies include anti-β-actin (Abcam, ab13822, 1:3000), anti-E-cadherin (BD, 610182, 1:200 for immunostaining, 1:1000 for Western blotting), anti-E-cadherin (Abcam, ab11512, Decma-1, 1:200), anti-G3BP2 (Sigma-Aldrich , HPA018425, 1:200, 1:1000), anti-Fibronectin (Sigma-Aldrich, F3648, 1:200), anti-Integrin α6 (Millipore, MAB1378, NKI-GoH3, 1:200), anti-human Laminin V (Chemicon, D4B5, 1:200), anti-mouse Laminin V (kind gift from M. Aumailley, 1:1000), anti-Twist1 (Santa Cruz, ab50887, Twist2C1a, 1:100, 1:1000), Rabbit anti-Twist1 (Sigma-Aldrich, T6451, 1:1000), 5b7 mouse anti-Twist1 hybridoma cell line (1:1000), anti-YAP1 (Santa Cruz, H-125, 1:100). AIIB2 hybridoma supernatant was used for β1 integrin blocking experiments (Developmental Studies Hybridoma Bank, 1:1000). Secondary fluorescent antibodies used include anti-mouse, anti-rat, and anti-rabbit conjugated with Alexafluor 488, 546, and 647 (Life Technologies). Secondary horseradish peroxidase (HRP) conjugated antibodies used include anti-mouse, anti-rabbit, and anti-chicken (Jackson Immunoresearch).
Wei S.C., Fattet L., Tsai J.H., Guo Y., Pai V.H., Majeski H.E., Chen A.C., Sah R.L., Taylor S.S., Engler A.J, & Yang J. (2015). Matrix stiffness drives Epithelial-Mesenchymal Transition and tumour metastasis through a TWIST1-G3BP2 mechanotransduction pathway. Nature cell biology, 17(5), 678-688.
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10

Immunoblotting and Immunofluorescence of YAP

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Primary antibodies used were: rabbit YAP (H-125, Santa Cruz, sc-15407; 1:200IF, 1:1000 IB), mouse YAP (63.7; Santa Cruz, sc-101199; 1:200 IF, 1:1000 IB), rabbit p-YAP (Cell Signaling Technology, 4911; 1:1000 IB). Samples were imaged with a Leica SP5 confocal microscope using a 63× oil immersion objective and processed using Adobe Photoshop. Fixation and cell culture quantification was carried out as previously described (Fletcher et al., 2015 (link)).
Elbediwy A., Vincent-Mistiaen Z.I., Spencer-Dene B., Stone R.K., Boeing S., Wculek S.K., Cordero J., Tan E.H., Ridgway R., Brunton V.G., Sahai E., Gerhardt H., Behrens A., Malanchi I., Sansom O.J, & Thompson B.J. (2016). Integrin signalling regulates YAP and TAZ to control skin homeostasis. Development (Cambridge, England), 143(10), 1674-1687.
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