I am trying to detect glycoproteins in SDS-PAGE gels using schiff's reagent. However my control (non glycosylated) protein is also being stained pink. I am following the Himedia protocol for the staining process. I am using Himedia S070 stain and Himedia S073 periodic acid. Can anyone point me to a working protocol?
Hi Dipayan,
The protocol for staining glycoproteins in SDS -PAGE should include the following steps;
1.Fix the gels using fixative solution (for eg., 50%Methanol)
2. Gently wash them using 3 to 5% acetic acid
3. Transfer the gels into oxidizing solution 10 to 15 mins (Periodic acid)
4. Wash with 3 to 5% acetic acid
5. Stain with Schiff' reagent.
6. Transfer the gels into reducing solution (Sodium metabisulphite)
7. Stop the reaction by transferring the gels into 3-5% acetic acid.
As mentioned in your query (Himedia), Try S074 (Basic fuschin) instead of S070 (Pararosaniline) along with S073.
S073 - Oxidizing reagent
S074 - Schiff reagent (Basic fuschin and Sodium metabisulphate)
Alternatively consider using Pierce Glycoprotein staining kit. Please find the link below.
All the best for your efforts.
https://assets.thermofisher.com/TFS-Assets/LSG/manuals/MAN0011387_Pierce_Glycoprotein_Stain_UG.pdf
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