Glycoprofile III

The classical method for in-gel carbohydrate detection uses Periodic Acid/Schiff reagent (PAS). It has a detection limit of 25-100 ng of carbohydrate (Product Code GlycoPro). PAS staining of glycoproteins is very selective, but lacks the sensitivity of fluorescent detection (5-25 ng of carbohydrate). The limit of detection varies with the glycoprotein and the degree and type of glycosylation. Typical detection limits observed with the ProteoProfile PTM Marker are 150 ng of ovalbumin (5 ng of carbohydrate) and 150 ng of RNase B (30 ng carbohydrate). The detection limits are 5-10 times lower than those observed with the Periodic Acid/Schiff reagent.

GlycoProfile III can also be used on PVDF membranes. After gel electrophoresis, the proteins are transferred to a PVDF membrane using standard procedures.

Sufficient reagents are supplied for 10 minigels

Components:
Oxidation Reagent
Glycoprotein Staining Reagent
Staining Buffer
ProteoProfile PTM Marker

• Designed as both a positive and negative control for SDS-PAGE gels and Western blots of proteins with post-translation modifications.

Figure 1: PTM Marker (2 ml of a 6-fold dilution), containing glycosylated and nonglycosylated proteins, was separated by electrophoresis on a 4-20% SDS-PAGE gel. The gel was stained for glycoproteins with GlycoProfile III (left), imaged, and then stained for total protein with EZBlue Gel Staining Reagent (right). The glycoproteins appear as bright fluorescent bands. Each band represents approximately 300 ng of protein.

Figure 2: Mouse IgG and rabbit IgG on a 4-20% SDS-PAGE gel and stained in the same manner as the gel in Figure 1. The IgG heavy chains, which are glycosylated, react strongly with the fluorescent detection reagent. 2.5 µg of protein was applied to each lane.