Enhanced detection of glycoproteins in polyacrylamide gels.
A highly sensitive and simple method to enhance detection of glycoproteins resolved by either one- or two-dimensional polyacrylamide gel electrophoresis is described. The method is a modification of the procedure described by D. Fargeaud et al. (D. Fargeaud, J. C. Benoit, F. Kato, and G. Chappuis (1984) Arch. Virol. 80, 69-82) that uses concanavalin A conjugated with fluorescein isothyocyanate to detect the carbohydrate moiety of glycoproteins. Briefly, the electrophoresed gel is exposed to the fluorescent lectin, thoroughly washed, and sequentially transferred to 50% methanol in deionized water and to absolute methanol. The result is an abrupt dehydration of the gel which turns evenly white and stiff. At least a twofold enhancement of fluorescence is obtained as detected by exposing the treated gel to an appropriate uv source. The sensitivity of the procedure allows us to detect purified immunoglobulin molecules by their carbohydrate content in the range of 0.2 microgram of total protein. The specificity of the detection is demonstrated by a comparison with the corresponding polypeptide profile obtained by silver nitrate staining of the gel.
Muñoz
G
G
Laboratorio de Genética Molecular, Facultad de Ciencias Básicas y Matemáticas, Universidad Católica de Valparaíso, Chile.
Marshall
S
S
Cabrera
M
M
Horvat
A
A
D004591:Electrophoresis, Polyacrylamide Gel; D006023:Glycoproteins / Q000032:analysis*; D006801:Humans; D007074:Immunoglobulin G / Q000032:analysis; D000432:Methanol; D013050:Spectrometry, Fluorescence
D006023:Glycoproteins; D007074:Immunoglobulin G; D000432:Methanol