High-performance liquid chromatographic determination of cyclic 3',5'-AMP with fluorescence detection. Vasoactive intestinal peptide-induced modification of its concentration in neuroblastoma cells.
The efficiency of ion-pair reversed-phase HPLC on a Vydac C18 column with 50 mM ammonium acetate (pH 4.75)-methanol-acetonitrile (88:9:3, v/v/v) as the mobile phase with isocratic separation and fluorescence detection for the determination of cAMP in cellular extracts was evaluated. This method was compared with a radioimmunoassay technique in terms of linearity, reproducibility and sensitivity. No interactions with other nucleotides such as AMP, ADP, ATP and cGMP were observed. Application to the measurement of cAMP modifications was studied in a neuroblastoma cell line: LA-N-2 cells stimulated by a neuropeptide, vasoactive intestinal peptide.
Laboratoire de Chimie Thérapeutique, Faculté des Sciences Pharmaceutiques et Biologiques, Lille, France.
J F
JF
Goossens
F
F
Leroux
N
N
Pommery
J
J
Pommery
J P
JP
Hénichart
D000085:Acetates; D002851:Chromatography, High Pressure Liquid / Q000379:methods* / Q000706:statistics & numerical data; D000242:Cyclic AMP / Q000032:analysis* / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D009447:Neuroblastoma / Q000378:metabolism*; D011863:Radioimmunoassay; D012680:Sensitivity and Specificity; D014407:Tumor Cells, Cultured; D014660:Vasoactive Intestinal Peptide / Q000494:pharmacology*
D000085:Acetates; D014660:Vasoactive Intestinal Peptide; D000242:Cyclic AMP; C018824:ammonium acetate