How many times can the same affinity column in the GlycoLink Immobilization Kit be used for purification?
The stability of the immobilized protein and the type of elution buffer used determines how many times a column can be reused. Typically, the columns can be reused at least five times without significant loss in performance.
Find additional tips, troubleshooting help, and resources within our Protein Purification and Isolation Support Center.
Will Hydrazide UltraLink Resins in the GlycoLink Immobilization kit couple chicken IgY?
Yes. Chicken antibodies have a total of 4 N-linked glycosylation sites and will couple to the resin.
Find additional tips, troubleshooting help, and resources within our Protein Purification and Isolation Support Center.
Can the Hydrazide UltraLink Resin in the GlycoLink Immobilization kit couple all types of antibodies?
The resin will couple any antibody that is adequately glycosylated. Although monoclonal antibodies are typically under-glycosylated, many have been effectively immobilized to this resin. To determine if a monoclonal antibody is glycosylated use the Thermo Scientific Glycoprotein Carbohydrate Estimation Kit (Cat. No. 23260).
Find additional tips, troubleshooting help, and resources within our Protein Purification and Isolation Support Center.
When oxidizing with sodium meta-periodate, is it possible to over-oxidize the antibody when using the GlycoLink Immobilization Kit?
If too much sodium meta-periodate is used or the sample is incubated for too long the oxidation can cause the protein to breakdown or favor an amine linkage to the resin. Furthermore, if used in excess, methionine residues might become oxidized. Typically, using 10 mM sodium meta-periodate for 30 minutes will oxidize only the carbohydrates and does not affect protein function.
Find additional tips, troubleshooting help, and resources within our Protein Purification and Isolation Support Center.
How do I oxidize a glycoprotein or antibody when using the GlycoLink Immobilization Kit?
The antibody or other glycoprotein is first diluted or desalted into the coupling buffer and transferred to a vial containing sodium meta-periodate at a final concentration of 10 mM (1 mM is sufficient for sialic acid residues). The mixture is incubated for 30 minutes at room temperature in the dark. Sodium meta-periodate concentration can be increased to 25mM if oxidation is not sufficient at 10 mM. Care must be taken not to over-oxidize the glycoprotein.
Find additional tips, troubleshooting help, and resources within our Protein Purification and Isolation Support Center.