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Cleanup of N-labelled glycans Cleanup of N-labelled glycans

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Micro-SPE Techniques (MiniSpin columns, 96-Well Spin plates)


Cleanup of N-labelled glycans in NEB Rapid PNGase F buffer from 2AB, or procainamide reagent, NaCNBH4, DMSO, and acetic acid labelling solution.

With thanks to Paula Magnelli, et al., New England Biolabs

Note: These loading volumes are high but the cartridges are on a vacuum manifold so the excess liquid is pulled through the cartridges.

HILIC cleanup (vacuum manifold) of MicroSpin HILIC (SEM HIL):
  • Condition SEM-HIL HILIC columns with 350µl Acetonitrile.
  • Add 350µl, 50mM ammonium formate, pH 4.4. Repeat two times.
  • Equilibrate with 350µl of a solution 90% Acetonitrile/10% 50mM ammonium formate, pH 4.4. Repeat two times
  • Bring glycan labeling reaction to 87-85% acetonitrile, mix well. Total volume 100-200µl
  • Load sample slowly into an equilibrated SPE column.
  • Wash with 350µl of a solution 90% Acetonitrile/10% 50mM ammonium formate, pH 4.4. Repeat five times
  • Spin at 3000 rpm 1 min to dry.
  • Elute with 50µl of 50mM ammonium formate, pH4.4: Let it enter by gravity, incubate for 2min. Spin 3000 rpm 2 min.

    Overview of All Chemistries and Configurations
    General Operating Instructions:
    General HILIC: Hydrophilic Interaction and Detergent Removal Instructions


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    Last Updated: 01/15/18
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