Glycoprotein Standards Set for Pierce™ Glycoprotein Carbohydrate Estimation Kit - FAQs

Yes. Reducing agents interfere with this assay. Please see the list of compatibility of other commonly used reagents below: Compatible: EDTA, Urea (<1.0M), Guanidine HCl (<1.0M), DMSO (<10%), PBS Not Compatible: Reducing Agents, Tris, Glycerol

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Most commonly used detergents do not interfere; however, detergents with sugar derivatives, such as octyl-b-glusoside (OG) and octyl-b-thioglucopyranoside (OTG), can contribute to the signal, resulting in an erroneously high percent carbohydrate value.

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Glycoprotein Carbohydrate Estimation Kit estimates the carbohydrate content of glycoproteins in solution; however, the Thermo Scientific Glycoprotein Stain directly stains oxidized glycoprotein on gels and nitrocellulose membranes and does not estimate carbohydrate content.

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The kit is designed for purified proteins but can also estimate the w/w percentage of carbohydrate in total protein.

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Yes. Free sugars form aldehydes when oxidized, which react with the Glycoprotein Detection Reagent.

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Yes. The purple solution produced when oxidized carbohydrates react with the Aldehyde Detection Reagent can be measured at 530 nm and 560 nm.

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The kit contains sufficient reagents for processing 250 microplate assays or 60 test tube assays and includes negative and positive controls.

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The assay's estimation range is 3.2% (ovalbumin) to 41.4% (a1-acid glycoprotein). The protein concentration required for the assay is based on the percent carbohydrate (w/w). Highly glycosylated proteins are assayed at 0.25mg/mL. Less glycosylated proteins are assayed at 2.5 mg/mL. Researchers with unknown samples should test them at both concentrations to ensure the sample is within the absorbance range for the standard curve.

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The results produced by this assay are an approximation of carbohydrate content, because the assay detects sialic acid, galactose and mannose. Therefore, sialic acid-rich glycoproteins will produce a higher absorbance than glycoproteins with a similar percentage of glycosylation but with fewer sialic acid residues.

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Glycoprotein is oxidized with sodium meta-periodate, which forms aldehydes. When reacted with aldehydes, the proprietary Aldehyde Detection Reagent becomes purple with an absorption maximum at 550 nm. The color intensity is proportional to the percentage of carbohydrate (aldehyde) component in the glycoproteins.

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