Ixodes scapularis Say, 1821 larvae were fed on mice and allowed to molt under laboratory conditions. A liquid chromatography-tandem mass spectrometry-based proteomic study was conducted to identify the type of mammalian proteins present in the derived nymphal ticks at different time intervals after molting. Albumin was present for 85 d; transferrin was present for 29 d; and, more importantly, hemoglobin remained detectable for up to 309 d postmolting. Peptides of actin, keratin, and tubulin are highly similar between mouse and tick, and therefore, unambiguous assignment of these proteins to different species was not possible. Establishing a time line for the persistence of hemoglobin, one of the most abundant blood proteins, at detectable levels in ticks after the bloodmeal and molting advances our efforts to use this protein to identify the host species.