Abstract
A sensitive enzyme-linked, indirect immunosorbent assay (ELISA) for the determination of guanosine 3′,5′-cyclic monophosphate (cGMP) in glial cells is described. The assay uses an antiserum produced against succinylated cGMP and is based on the competition between endogenous cGMP and a fixed amount of immobilized antigen. The antibody exhibited a high degree of specificity with negligible cross reactivity with other nucleotides or related compounds. The standard curve, linearized using a logit–log transformation, had an operating range of 1 fmol/50 μl to 5 pmol/50 μl. The sensitivity of the assay was significantly increased by acetylation of standards and samples. Recoveries of cGMP from samples of cultured cells ranged from 85% to 105% with a mean recovery of 97% ± 7%. Levels of cGMP measured with the ELISA were in agreement with the corresponding values obtained using radioimmunoassay. The present method provides for a cheap, sensitive and simple alternative to the commercially available cGMP assays.
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Special issue dedicated to Dr. Lawrence F. Eng.
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Wellard, J., Blind, B. & Hamprecht, B. An Enzyme-Linked Immunosorbent Assay for the Rapid Quantification of Intracellular and Extracellular Guanosine 3′,5′-cyclic Monophosphate in Cultured Glial Cells. Neurochem Res 29, 2177–2187 (2004). https://doi.org/10.1007/s11064-004-8675-x
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DOI: https://doi.org/10.1007/s11064-004-8675-x