Calculation of the spectroscopic correction factor.

Abbreviations: MUNANA, 2′-(4-methylumbelliferyl)-α-D-N-acetylneuraminic acid; 4-MU, 4-methylumbelliferone; RFU, relative fluorescence units.

a

Fluorescence of MUNANA alone in enzyme buffer at 1.95–2000 µM concentrations (shown in column 1).

b

Fluorescence of 4-MU alone in enzyme buffer at 19 µM concentration. Mean (± SD) 4-MU fluorescence in enzyme buffer was 10222±90 RFU.

c

Fluorescence of 4-MU in enzyme buffer at 19 µM concentration in the presence of 1.95–2000 µM MUNANA concentrations (shown in column 1).

d

Values (in RFU) of MUNANA fluorescence (column 2) subtracted from the values (in RFU) obtained in the mixture of 4-MU and MUNANA (column 4).

e

Correction factor calculated as the ratio of 4-MU fluorescence (in RFU) in the mixture with MUNANA substrate (column 5) and 4-MU fluorescence (in RFU) alone (mean value from column 3).