Determination of neuraminidase-susceptible and total N-acetylneuraminic acid in cells by selected ion monitoring. |
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Authors: | J Roboz R Suzuki J G Bekesi |
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Affiliation: | Department of Neoplastic Diseases, The Mount Sinal School of Medicine, City University of New York, New York, N. Y. 10029 USA |
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Abstract: | N-acetylneuraminic acid (NANA) is released from glycoproteins by neuraminidase or acid hydrolysis and quantified by monitoring the protonated molecular ions of fully silylated NANA () and neuraminic acid β-methylglycoside (internal standard, ) in a gas chromatograph—mass spectrometer system using isobutane ionization. Detection limit is 200 pg (0.6 pmol, underivatized weight) of NANA injected. In biological samples 5 ng (15 pmol) of NANA can be detected in 50 μl of hydrolysate. Only 1 to 50 μl of hydrolysate is needed, sample preparation is simple, NANA is positively identified in every analysis, 2-deoxy carbohydrates and other sialic acids do not interfere, only free NANA is determined, and the internal standard increases reliability. The NANA content of neuraminidase-treated human leukemic cells was on the order of cells. NANA was quantified using as few as 5 × 104 cells, in contrast to the conventional colorimetric (thiobarbituric) technique which requires 2.5 × 107 cells. |
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