2-AB Labeling Kits

Product Description 2-AB (2-aminobenzamide) is one of the most widely used fluorescent labels for glycosylation analysis. The 2-AB Labeling Kit includes all the reagents needed to label up to 30 glycan samples. Kits are available with two different reducing agent types: sodium cyanoborohydride or nontoxic 2-picoline borane. Traditional 2-AA and 2-AB labeling kits use sodium cyanoborohydride as a reducing agent during glycan labeling. This reagent is toxic, so a fume hood should be used during handling. To comply with emerging health and safety regulations, we are now replacing these kits with our new VP Glycan Kit, which uses picoline borane, a safer reducing agent. The 2-AB Labeling Kit contains two sets of the following reagents (15 samples per kit) in glass ampoules sealed under pure nitrogen: A492565-01-24T 2-Aminobenzamide (2-AB dye) Dimethyl Sulfoxide (DMSO) Acetic acid Sodium cyanoborohydride A492565-02-24T/A492565-03-96T 2-Aminobenzamide (2-AB dye) Dimethyl sulfoxide (DMSO) / acetic acid solution 2-Methylpyridineborane Number of samples: One 2-AB labeling kit contains reagents and each kit can label up to 30 individual samples for analysis Dye purity: >99% by HPLC Molecular weight: 137 Lambda-ex: 250 nm Lambda-em : 428 nm Sample amounts per sample ranged from 25 pmol to 25 nmol glycans. Appropriate samples can label any purified glycan with free reducing ends. No detectable ( Labeling efficiency is typically > 85 % (depending on the sample). Label selectivity is essentially stoichiometric labeling. Program 1. Purification of glycans: purify glycans from proteins, salts and detergents. 2. Transfer the sample to the reaction vial: For glycan pools obtained from typical glycoproteins, the sample volume should be between 100 picomolar and 50 nanomolar. In subsequent HPLC analysis, single pure glycans down to 5 picomolar can be labeled and detected. Suitable reaction vials include small polypropylene microcentrifuge tubes and PCR work tubes. 3. Dry the sample: If the sample volume exceeds 10 μL, dry the sample. If the sample needs to be dried, it should be dried using a centrifugal evaporator. If this is not possible, freeze-drying (lyophilization) can be used sparingly (especially to ensure that the sample dries to a small, dense mass at the bottom of the vial). Do not expose samples to high temperature (>28°C) or extreme pH conditions, as these conditions can lead to acid-catalyzed loss of sialic acid (high temperature, low pH) or glycan reducing end epimerization (at high pH) conditions). After the sample has dried, redissolve the glycans in 10 μL of water. 4. Prepare the DMSO-acetic acid mixture: Add 150 µL of glacial acetic acid to the DMSO vial and mix by pipette motion. 5. Add the dye: Add the DMSO-acetic acid mixture to a bottle of 2-AB (2-aminobenzamic acid) dye and mix until the dye dissolves. 6. Add the reducing agent: Add the dissolved dye to a vial of sodium cyanoborohydride or picoline borane and mix with a pipette until the reducing agent is completely dissolved to make the final labeling reagent. 7. Add labeling reagent to samples: Add labeling reagent to each dried glycan sample, cap the microtube, and mix well. 8. Incubation: Incubate the reaction vial in a heating block, sand tray, or drying oven set to 65°C for 3 hours. In most cases, the incubation time can be shortened to 2 hours or extended to 4 hours without significantly altering the results of the labeling reaction. 9. Centrifugation and cooling: After the incubation period, remove the samples, centrifuge the microtubes briefly, and allow them to cool completely to room temperature. 10. Sample clean-up: It is recommended to clean the labeled sample to remove excess dye and other labeling reagents. Purification can be achieved with a T1 or S-filter

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