First successful detection of oestrogen, progesterone and testosterone in multiple human hard tissues, and their use as potential biomarkers of pregnancy

The sex steroid hormones oestrogen, progesterone, and testosterone have never been detected in modern or archaeological human skeletal tissues using enzyme-linked immunosorbent assay (ELISA) analysis, and there are no standard protocols for their extraction. As progesterone is a biomarker of pregnancy in living individuals, its detection in skeletal remains would substantially improve the visibility of pregnant individuals in the archaeological record and furnish a novel means of exploring female life histories in the past. The present study demonstrates that oestrogen, progesterone and testosterone can be detected in the hard tissues of ten individuals of known sex dating from the 1st to 19th centuries CE and evaluates their potential as biomarkers of pregnancy. The cohort comprised seven females of varied parity status and three males. A novel ELISA methodology was developed for hormone extraction from prepared tissue samples of bone, dentine, enamel, root from second and third permanent molars, and dental calculus (n = 74). Oestrogen, progesterone and testosterone concentrations were measurable in bone, dentine, enamel, and root samples, whereas only progesterone and testosterone were detected in dental calculus. Elevated progesterone concentrations were detected in the bone and tooth structures of one pregnant female, all individuals with in utero pregnancies had undetectable testosterone, and those associated with fetal remains presented elevated progesterone levels in dental calculus. Our findings demonstrate the feasibility and significant potential of the ELISA method for the detection of sex hormones in human skeletal remains to examine the reproductive histories of past populations.