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The present study is a preliminary evaluation of the potential for an elevation of relaxin to clarify an elevation of progesterone in wild bottlenose dolphins as pregnancy-related since relatively high progesterone in a single-sample analysis is not necessarily pregnancy-specific. Blood samples were collected from adult female bottlenose dolphins in estuaries of Charleston, South Carolina, and the Indian River Lagoon, Florida. Serum progesterone and relaxin concentrations were analyzed in the same samples (n = 74) with respective radioimmunoassays (RIAs). Diagnosis of pregnancy was based on concentrations of progesterone and relaxin > 6.0 and > 11.2 ng/mL, respectively, whereas non-pregnancy was based on respective hormone concentrations less than or equal to baseline concentrations. Progesterone was observed at elevated concentrations considered diagnostic of pregnancy in 20 and relaxin in 12 bottlenose dolphins. In a contingency table analysis with progesterone as the conventional standard for pregnancy diagnosis, there was a 60% positive correspondence with relaxin (12 relaxin-out of 20 progesterone-diagnosed pregnancies) and 40% negative correspondence (8 relaxin-diagnosed nonpregnancies out of 20 progesterone-diagnosed pregnancies). Kappa analysis indicated good agreement (. = 74%) between relaxin-and progesterone-diagnosed pregnancies. Contingency table results were verified based on known pregnancies of nine cow-calf pairs observed through photo-identification subsequent to health assessments. Concordance between hormone diagnosed and observed pregnancies was 9 of 9 (100%) for progesterone and 7 of 9 (78%) for relaxin. Hence, preliminary results indicated that relaxin may have limited value for pregnancy diagnosis when used alone. However, in combination with an elevation of progesterone in a single-sample analysis, a pregnancy-specific elevation of relaxin can clarify that progesterone is pregnancy-related and, thus, confirm the diagnosis of pregnancy in wild bottlenose dolphins.