Analysis of glycoproteins in cell culture supernatants using a lectin immunosensor technique.
A method based on a surface plasmon resonance technique for detection of changes in concentration and glycosylation of proteins in cell culture supernatant is described. The method was used to analyze alpha(1)-acid glycoprotein (AGP) produced by a human hepatoma cell line (HepG2). Cell culture supernatant was injected to a BIACORE 2000 instrument and AGP was captured on the sensor chip by immobilized antibodies. The captured glycoprotein was then analyzed for content of carbohydrate epitopes using three different lectins, Aleuria aurantia lectin (AAL), Sambucus nigra agglutinin (SNA), and Triticum vulgaris agglutinin (wheat germ agglutinin, WGA). The method was used to analyze changes in concentration and glycosylation of AGP produced by HepG2 cells grown with or without three different cytokines, interleukin-1 beta (IL-1 beta), interleukin-6 (IL-6), and transforming growth factor beta-1 (TGF beta(1)). Using the described method it was shown that when HepG2 cells were grown in the presence of IL-6 both AGP concentration and fucosylation increased. When HepG2 cells instead were grown in the presence of TGF beta(1) AGP fucosylation increased whereas AGP concentration decreased.
Liljeblad
Mathias
M
Department of Biomedicine and Surgery, Division of Clinical Chemistry, Linköping University, SE-581 85, Linköping, Sweden.
Lundblad
Arne
A
Påhlsson
Peter
P
D015374:Biosensing Techniques / Q000379:methods*; D006023:Glycoproteins / Q000032:analysis* / Q000378:metabolism; D006031:Glycosylation; D006801:Humans; D007118:Immunoassay; D015850:Interleukin-6 / Q000494:pharmacology; D037102:Lectins / Q000378:metabolism*; D009961:Orosomucoid / Q000032:analysis* / Q000276:immunology; D012680:Sensitivity and Specificity; D020349:Surface Plasmon Resonance; D016212:Transforming Growth Factor beta / Q000494:pharmacology; D053773:Transforming Growth Factor beta1; D014407:Tumor Cells, Cultured
D006023:Glycoproteins; D015850:Interleukin-6; D037102:Lectins; D009961:Orosomucoid; C508960:TGFB1 protein, human; D016212:Transforming Growth Factor beta; D053773:Transforming Growth Factor beta1