Establishment of a reliable extraction method of estrone sulfate from bovine plasma for detection of the peripheral level during the regular estrous cycle by radioimmunoassay

A simple extraction and assay technique of estrone sulfate in bovine blood was developed with the object of detecting the peripheral level of estrone sulfate in a normal estrous cycle or in early pregnancy. Estrone sulfate in bovine plasma was extracted with a small reversed phase cartridge. The steroid conjugate retained in the cartridge was eluted with 40% (v/v) methanol. Estrone sulfate was separately recovered from other steroids by the stepwise increase in methanol concentration in the elution solvent. The recoveries of estrone sulfate eluted with 40% methanol were more than 90%, irrespective of the applied plasma volume. The concentration measured by radioimmunoassay with the eluent of 40% methanol was consistent for plasma extraction volumes of 0.5–2.0 ml. The change of estrone sulfate in bovine peripheral plasma during the regular estrous cycle was determined with a small reversed phase cartridge for extraction and 40% methanol for elution. The change in estrone sulfate was found to be similar to the change of estrone and estradiol-17β. The concentration of estrone sulfate was not higher than that of both estrogens in cattle.