Total protein was extracted from 5-day-old seedlings that had been exposed to white light for 3 h, transferred to darkness for 21 h, and exposed to continuous red light for 4 days. Protein concentrations were determined by the Bradford assay (Bio-Rad). Thirty micrograms of protein extract were separated by SDS-PAGE, transferred onto polyvinylidene fluoride (PVDF) membranes, and treated with horseradish peroxidase (HRP)-conjugated antibodies against GFP (anti-GFP-HRP, 1:10 000, Miltenyi Biotec) or anti-tubulin (mouse monoclonal, 1:10 000, Sigma-Aldrich) and HRP-conjugated anti-mouse antibodies (rabbit polyclonal, 1:10 000, Abcam). The blots were visualized using the Western Lightning Plus Enhanced Chemiluminescence kit (PerkinElmer) and the X-Doc System (Bio-Rad). Precision Plus Protein Dual Color Standards (Bio-Rad) were used as protein size markers.
Western lightning plus enhanced chemiluminescence kit
Manufactured by PerkinElmer
The Western Lightning Plus Enhanced Chemiluminescence kit is a laboratory reagent used for the detection and quantification of proteins in Western blot analysis. The kit includes reagents that generate a chemiluminescent signal upon interaction with the target protein, allowing for sensitive and specific detection.
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Lab products found in correlation
2 protocols using western lightning plus enhanced chemiluminescence kit
1
Analysis of Protein Expression in Arabidopsis
2
Immunoblot Detection of GFP-Tagged Proteins
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Proteins were extracted from plant material with extraction buffer (150 mM Tris-HCl, pH 7.5, 150 mM NaCl, 10% [v/v] glycerol, 10 mM EDTA, 1 mM sodium molybdate, 1 mM NaF, 10 mM dithiothreitol, 1% [v/v]) NP-40, 0.5% [v/v] polyvinylpolypyrrolidone, protease inhibitor). Total protein content was determined with the Qubit protein assay kit (Invitrogen). Total proteins were separated on 4-12% gradient Mini-PROTEAN stain-free TGX gels (Bio-Rad) and transferred to a polyvinylidene difluoride membrane. Isolated proteins were detected by immunoblot analysis with monoclonal rabbit GFP-HRP antibody (Abcam) at a 1:2000 dilution and the signal was identified through chemiluminescent substrates from the Western Lightning® Plus Enhanced Chemiluminescence kit (PerkinElmer) under a ChemiDoc Imaging system (Bio-Rad).
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