Recombinant bovine somatotropin (rbST) is used in dairy cattle to enhance milk production. Despite the ban on this hormone in some countries, especially in Europe, there is so far no method available for the direct detection of rbST either in milk or in plasma. An analytical strategy has been developed to analyze rbST in plasma, including a purification procedure based on a precipitation with ammonium sulphate, followed by a solid-phase extraction (SPE)-based clean-up on C4 sorbent and precipitation with cold methanol. The hormone was then digested with trypsin and analyzed by liquid chromatography/high-resolution mass spectrometry (LC/HRMSn) on a linear ion trap coupled with an Orbitrap. The tryptic N-terminal peptide, specific to the difference between the endogenous and recombinant form of the somatotropin, was fragmented and product ions were analyzed at high mass resolution. Applying this approach to goat plasma allowed the direct detection of 10 ng mL(-1) of rbST in fortified samples. It also showed the presence of rbST in plasma collected from a goat treated with the hormone, even 2 days after administration. These results are of a great interest in the field of somatotropin control and undoubtedly constitute a first step in the development of a method for the detection of rbST not only in bovine plasma, but also in other biological matrices such as milk.