2.6.2. Sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS-PAGE)
This protocol is extracted from research article:
Effect of ultrasonic pre-treatment on Ara h 1 in peanut sprouts
Ultrason Sonochem, May 25, 2021; DOI: 10.1016/j.ultsonch.2021.105607

SDS-PAGE was used to separate proteins extracted from the degreased powder using an SDS Kit obtained from Solarbio (Sinopharm Chemical Reagent Co., Ltd, Beijing, China). The SDS-PAGE method was modified from that described in Meng et al. [21] and used a Mini Protein Tetra System (BioRad, Hercules, CA, USA). Each electrophoresis sample (2.0 mg) was placed in a 1.5-mL centrifuge tube. Soluble protein extracts were mixed with an equal volume of sample buffer containing 1% β-mercaptoethanol. The mixture was boiled for 5–10 min for denaturation. Electrophoresis was performed on 13% acrylamide gels for 40 min at 80 V, followed by 3 h at 120 V. Protein marker standards were used to identify and estimate the major allergens and their molecular masses. To quantify the peanut major allergen Ara h 1, gels were scanned and analyzed with a Molecular Imager (AI600UV, GE Analytical Instruments, USA). The major allergens and their relative composition were calculated based on the band intensity and total area of their subunits.

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