The quantitative determination of 2'-deoxycytidine-5'-triphosphate in cell extracts by radioimmunoassay.
A radioimmunoassay (RIA) capable of quantitating dCTP in femtomolar amounts in cell extracts has been developed, and applied to human fibroblast cell lines and L5178Y mouse lymphoma lines. Cross reactivity of the antibody with CTP, though low (2.7%) has necessitated pre-RIA removal of CTP by either boronate affinity gel chromatography or sodium periodate oxidation. Fractions from the boronate gel column or aliquots of NaIO4-treated cell extract are quantitated directly by the RIA. Recovery of extracted dCTP standard taken through the entire procedure is quantitative and results are reproducible. Due to the high sensitivity of the quantitation step, dCTP can be accurately measured in relatively small numbers of cells--about 10(4) cells.
D000818:Animals; D002846:Chromatography, Affinity; D003429:Cross Reactions; D003570:Cytidine Triphosphate / Q000276:immunology / Q000302:isolation & purification; D003845:Deoxycytosine Nucleotides / Q000032:analysis* / Q000276:immunology; D005347:Fibroblasts / Q000032:analysis; D006801:Humans; D007940:Leukemia L5178 / Q000378:metabolism; D051379:Mice; D008832:Microchemistry; D010504:Periodic Acid; D011863:Radioimmunoassay / Q000379:methods*
D003845:Deoxycytosine Nucleotides; D010504:Periodic Acid; C024107:2'-deoxycytidine 5'-triphosphate; D003570:Cytidine Triphosphate; C009288:metaperiodate