Proteins were transferred to nitrocellulose membrane (GE Healthcare, AmershamTM HighbondTM-ECL) via wet western blotting at 4 °C and 30 V overnight. Membranes were blocked in TBS-Tween (0.1%)–Milk (5%) for 1 h at room temperature. For mCherry detection, membranes were incubated overnight with an anti-mCherry antibody (ab167453, dilution 1 : 2000; Abcam) followed by 1 h incubation with an anti-rabbit-horseradish peroxidase (HRP) antibody (dilution 1 : 10,000, Calbiochem). For GFP detection, membranes were incubated overnight with an anti-GFP-HRP antibody (130-091-833, dilution 1 : 1000, Miltenyi Biotec). For FLAG detection, membranes were incubated overnight with an anti-FLAG-HRP antibody (A8692, dilution 1 : 1000, Sigma). For SPX1 detection, membranes were incubated overnight with an anti-SPX1 antibody (dilution 1 : 1000, kind gift of Professor Mingguang Lei) followed by one hour incubation with an anti-rabbit-HRP antibody (dilution 1 : 10,000, Calbiochem). Antibodies were diluted in TBS-Tween (0.05%)–Milk (2.5%). Membranes were detected with SuperSignal™ West Femto Maximum Sensitivity Substrate (34095, Thermo Scientific™).
Anti gfp hrp antibody
Manufactured by Miltenyi Biotec
Sourced in United States
The Anti-GFP-HRP antibody is a detection reagent used to identify and visualize green fluorescent protein (GFP) in various applications. It is conjugated to horseradish peroxidase (HRP), which allows for sensitive detection through colorimetric or chemiluminescent methods.
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2 protocols using anti gfp hrp antibody
1
Western Blot Analysis of Protein Expression
2
GFP-Based Protein Detection Assay
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pRGA:GFP-RGA, 35S:EIN3-GFP, and 35S:HA-IAA1 (Silverstone et al., 2001 (link); Guo and Ecker, 2003 (link)) seeds were germinated in 2 mL liquid MS media in an orbital shaker at 100 rpm and the described compounds were added directly to the media 6 d after germination. After 2 h, 20 plants of the same size were collected and proteins were extracted as previously described (Fonseca and Solano, 2013 ). Samples were denatured, loaded on 9% sodium dodecyl sulphate–polyacrylamide gel electrophoresis (SDS–PAGE) gels, transferred to nitrocellulose membranes (Bio-Rad, Hercules, CA, USA), and incubated with anti-GFP-HRP antibody (Milteny Biotec, Bergisch Gladbach, Germany). Blots were developed using ECL (Pierce, Waltham, MA, USA).
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