OBJECTIVE To evaluate accuracy, user variability and impact of experience on the use of serum hCG and progesterone in women who have a pregnancy of unknown location (PUL's). METHODS This was a retrospective study. Presenting 1932 consecutive women to an Early Pregnancy Unit had a transvaginal scan. The location of the pregnancy could not be found in 189 women (Pregnancy of unknown location, PUL), and so blood was taken to measure serum hCG and progesterone at presentation and subsequently after 48 h, according to the protocol. All women were monitored at regular intervals until the final outcome was known, which was a failing PUL, a viable or failing intra-uterine pregnancy, an ectopic pregnancy or a persisting PUL. The final study group comprised 185 PUL, as four cases of persisting PUL were treated and excluded from the analysis. Five investigators assessed the hormonal data independently. The investigator's experience as defined by the number of years working in obstetrics and gynecology ranged from 2 to 15 years. Each investigator knew the women were clinically stable and that the scan result was consistent with a PUL, i.e. there were no signs of intra- or extra-uterine pregnancy, and there was no hemoperitoneum on TVS. When assessing the PUL's, each investigator was given the hormonal results at time 0 and 48 h for serum hCG and progesterone and asked to classify the PUL's as failing PUL's, immediately viable intra-uterine PUL's and ectopic PUL's. No other clinical information about the women was made available. RESULTS Complete data 185 women (89%): 102 failing PUL's, 63 immediately viable intra-uterine PUL's and 20 ectopic PUL's (total 185). The most experienced investigator obtained the best accuracy 163/185 (88.1%); not significantly different from those obtained by less experienced investigators (range 85.9-87.6%). Mean correct classification of failing PUL and immediately viable intra-uterine PUL's was 93% (range 89-95%); corresponding value for ectopic PUL's was 42% (range 25-60%). Agreement between observers for classification of failing PUL's and immediately viable intra-uterine PUL's was almost perfect (Cohen's kappa 0.86-0.90), whereas the value for ectopic PUL's group was fair to moderate (Cohen's kappa 0.39-0.67). All 5 investigators misdiagnosed same 35% of ectopic PUL's. CONCLUSIONS Serum hCG and progesterone levels at defined times can be used to predict the immediate viability of a PUL, but cannot be used reliably to predict its location. Clinical experience does not significantly improve the ability to assess PUL outcome.