Isolate glycosylated proteins from complex protein mixtures.

Glycosylation is a post-translational modification that plays an important role in biological functions, including immune regulation, inflammation, cell-to-cell adhesion, and cell signaling. We have developed two lectin-based Glycoprotein Isolation Kits, Concanavalin A (ConA) and Wheat Germ Agglutinin (WGA), that allow isolation of glycoproteins from complex protein mixtures, including serum, tissue and cultured cell lysates thus enabling downstream analysis.

ConA lectin, recognizes α-linked mannose and terminal glucose residues while WGA lectin, selectively binds to N-Acetyl glucosamine (GlcNAc) groups and to sialic acid.

Highlights

• High recovery – equivalent or greater glycoprotein recovery versus competitor kits and lectin resins
• Fast – glycoprotein purification in less than an hour
• Versatile – isolate glycoproteins from various sample types, e.g., human serum and cell lysate.
• Robust – lectin does not leach from resin when processing sample
• Convenient – complete kit offering lectin resins and spin columns with all necessary reagents
• Compatible with Bradford-based protein assays – dialysis or protein precipitation of recovered glycoproteins is not required prior to protein assay

Figure 1. Glycoprotein isolation from human serum and cell lysate – performance comparison of kits using WGA resin. CHO lysate and human serum samples were processed with the Pierce Glycoprotein Isolation Kit, WGA and with other commercially available WGA resins. An equivalent amount of total protein was applied to each resin. Eluted glycoprotein fractions were normalized by volume and resolved on 8-16% polyacrylamide gels. (a) Eluted glycoprotein fraction from applied CHO lysate (b) Eluted glycoprotein fraction from applied human serum.

Figure 2. Glycoprotein isolation from human serum and cell lysate – performance comparison of kits using ConA resin. CHO lysate and human serum samples were processed with the Pierce Glycoprotein Isolation Kit, ConA and with other commercially available ConA resins. An equivalent amount of total protein was applied to each resin. Eluted glycoprotein fractions were compared with ConA Resin boiled in SDS-PAGE Buffer to release lectins. All fractions were normalized by volume and resolved on 8-16% polyacrylamide gels. Gels were then silver stained. (a) Eluted glycoprotein fractions from applied CHO lysate (b) Eluted glycoprotein fractions from applied human serum. Arrows identify protein bands that result from ConA leaching from the resin during the elution process.

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