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Spa 896

Manufactured by Stressgen
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The SPA-896 is a laboratory equipment that measures the absorbance of light in liquid samples. It is designed to conduct spectrophotometric analysis, a widely used technique in various scientific disciplines to quantify the concentration of specific substances in a solution.

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Lab products found in correlation

Luciferase assay kit, by Promega (2 mentions) Nrf2 sc 722, by Santa Cruz Biotechnology (2 mentions) Kas tf110, by Stressgen (2 mentions) Pgl3 promoter plasmid, by Promega (1 mentions) Tricarbonyl dichlororuthenium 2 dimer tcdc, by Merck Group (1 mentions) Lycopene, by Merck Group (1 mentions) Vectastain abc reagent, by Vector Laboratories (1 mentions) Permount, by Thermo Fisher Scientific (1 mentions) Anti flag m2 fitc, by Merck Group (1 mentions) Mowiol 4 88, by Merck Group (1 mentions) Tcs sp2 confocal microscope, by Leica (1 mentions)

4 protocols using spa 896

1

Luciferase Assay for Nrf2 and NF-κB

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The p3xARE/Luc and NF-κB/Luc vectors were constructed by introducing the Nrf2 binding site or NF-κB binding site into the pGL3 promoter plasmid (Promega, Madison, WI, USA), respectively as described in a previous study (22) . Luciferase assay kits were purchased from Promega. Antibodies against HO-1 (SPA-896) and p65 (KAS-TF110) were purchased from Stressgen Biotechnologies (SB, San Diego, CA, USA). ICAM-1 (sc-7891), IκBα (sc-847), lamin B1 (sc-56143), α-tubulin (sc-53646) and Nrf2 (sc-722) antibodies were obtained from Santa Cruz Biotechnology (Santa Cruz, CA, USA). Bacteriallyderived TNFα was purchased from Calbiochem (San Diego, CA, USA). All other reagents, including lycopene and tricarbonyl dichlororuthenium (II) dimer (TCDC) were purchased from Sigma (St. Louis, MO, USA). The CO donor, TCDC, was activated by adding the compound to culture medium to liberate CO (20) .
Yang P.M., Chen H.Z., Huang Y.T., Hsieh C.W, & Wung B.S. (2017). Lycopene inhibits NF-κB activation and adhesion molecule expression through Nrf2-mediated heme oxygenase-1 in endothelial cells. International journal of molecular medicine, 39(6).
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2

Luciferase Assay for Cellular Stress Responses

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Materials. The p3xARE/Luc vector was constructed as described previously (16) . Luciferase assay kits were purchased from Promega Corporation (Madison, WI, USA). Antibodies against HO-1 (SPA-896) and p65 (KAS-TF110) were purchased from Stressgen Biotechnologies (San Diego, CA, USA). ICAM-1 (sc-7891), IκBα (sc-847), Lamin B1 (sc-56143), α-tubulin (sc-53646) and Nrf2 (sc-722) antibodies were obtained from Santa Cruz Biotechnology, Inc. (Santa Cruz, CA, USA). TrxR1 antibody (07-613) was purchased from Upstate (Charlottesville, vA, USA). IκBα and Nrf-2 antibodies were obtained from Santa Cruz Biotechnology, Inc. Bacterially derived TNF-α was purchased from Calbiochem (EMD Millipore; Billerica, MA, USA). All other reagents were purchased from Sigma-Aldrich (Merck KGaA, Darmstadt, Germany).
Chen L.G., Zhang Y.Q., Wu Z.Z., Hsieh C.W., Chu C.S, & Wung B.S. (2018). Peanut arachidin-1 enhances Nrf2-mediated protective mechanisms against TNF-α-induced ICAM-1 expression and NF-κB activation in endothelial cells. International journal of molecular medicine, 41(1).
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3

Immunohistochemistry for Heme Oxygenase-1

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Immunohistochemical staining was performed as previously described [8 (link)]. In brief, after deparaffination, blocking endogenous peroxidase activity and non-specific binding sites steps were carried out. Slides were then incubated with primary rabbit polyclonal anti-HO-1 antibody (dilution 1:100; SPA-896; Stressgen Bioreagents) overnight at 4 °C followed by incubation with diluted biotinylated secondary antibody and then incubation with VECTASTAIN ABC Reagent (Vector Laboratories, Inc., Newark, CA, USA). For negative controls, the primary antibody was omitted. Diaminobenzidine/H2O2 was used as a substrate for the immunoperoxidase reaction. Finally, they were lightly counterstained with hematoxylin and mounted with Permount (Fisher Scientific, Pittsburgh, PA, USA) for analysis by bright-field microscopy.
Mascaró M., Alonso E.G., Schweitzer K., Rabassa M.E., Carballido J.A., Ibarra A., Alonso E.N., Bermúdez V., Fernández Chavez L., Coló G.P., Ferronato M.J., Pichel P., Recio S., Clemente V., Fermento M.E., Facchinetti M.M, & Curino A.C. (2022). Hemoxygenase-1 Promotes Head and Neck Cancer Cell Viability. Antioxidants, 11(10), 2077.
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4

Immunofluorescence Analysis of HO-1 and FLAG Proteins

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To evaluate HO-1 expression by indirect immunofluorescence, cells were cultured on coverslips, and IIF was carried out as previously described, with a few modifications [8 (link),19 (link)]. Then, cells were incubated with a rabbit polyclonal anti-HO-1 (SPA-896; Stressgen Bioreagents) followed by an Alexa Fluor 488-conjugated secondary Ab (Molecular Probes). To evaluate FLAG expression in stably transfectants cells by direct immunofluorescence, cells were incubated with a mouse monoclonal anti-FLAG M2-FITC (F4049; Sigma Aldrich, Burlington, MA, USA) following the manufacturer’s instructions. Finally, cells were stained with DAPI and mounted on Mowiol® 4-88 (#81381, Sigma Aldrich). Cells were analyzed by using Nikon Eclipse E600 fluorescence microscope or Leica TCS SP2 confocal microscope.
Mascaró M., Alonso E.G., Schweitzer K., Rabassa M.E., Carballido J.A., Ibarra A., Alonso E.N., Bermúdez V., Fernández Chavez L., Coló G.P., Ferronato M.J., Pichel P., Recio S., Clemente V., Fermento M.E., Facchinetti M.M, & Curino A.C. (2022). Hemoxygenase-1 Promotes Head and Neck Cancer Cell Viability. Antioxidants, 11(10), 2077.
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