Supplementary Information

Each SiaFind™  detection reagent and enzyme is 6xHis-tagged (SP2302 and SP0502) or 8xHis-tagged (SP2602 and GE0701) at its N-terminus, and an anti-polyhistidine antibody or, in the case of the biotinylated version, a streptavidin conjugate can be used for detection. The 8xHis tag may be removed by FasTEV™ (Cat #GE0501), a TEV protease with enhanced stability and catalytic activity.

Each kit also includes specially formulated 5X binding buffers to ensure maximum reagent specificity and ease of use. For convenience, a pan-specific sialidase and associated 10X reaction buffer is included for efficient removal of sialic acid moieties to serve as controls for experiments.

Form and Storage

The SiaFind™ Lectenz® and reagents are supplied lyophilized in a storage buffer (50 mM EPPS, 200 mM NaCl, pH 7.5) and should be reconstituted in 100 μL molecular water grade to yield a 10 mg/mL solution (SP2302 and SP0502) or 1 mg/mL solution (SP2602). Concentration is determined by spectrophotometry using E1% 22.0. Once reconstituted, store at 4°C for up to 5 days or -20°C for up to 6 months. Aliquoting is recommended to avoid repeated freeze-thaw cycles.

The Pan Sialidase is supplied supplied lyophilized in a storage buffer (50 mM EPPS, 200 mM NaCl, pH 7.5) and should be reconstituted in 100 μL molecular water grade to yield a 50,000 units/mL solution. Once reconstituted, store at 4°C for up to 5 days or -20°C for up to 6 months. Aliquoting is recommended to avoid repeated freeze-thaw cycles.

All 5X buffers should be diluted to 1X with ultrapure water. For instance, to make 250 mL, add 50 mL of any 5X buffer to 200 mL water and mix by inversion.  All buffers may be stored at 4 to 25°C.

An optional α2,3-specific sialidase (GE0302) may be used along with kit SK0101 or SK0101B to maximize the capability of sialic acid linkage profiling.