Quantifying Testosterone and Corticosterone

Testosterone and corticosterone quantification was determined using Agilent’s Infinity II UHPLC in line with a 6495 triple quadrupole mass spectrometer and MassHunter workstation software (8.0.8.23.5). Briefly, DSRCT xenograft and PDX samples were homogenized using water containing internal standard (Cerilliant, T070) extracted with tert-butyl methyl ether (Sigma 34875), dried under nitrogen, and derivatized using hydroxylamine hydrochloride (Sigma 431362). The recovered ketoxime steroids were reconstituted in methanol/water (1:1 v/v) and injected into the Infinity II UHPLC. Ketoxime steroids were separated using a Chromolith reverse-phase column (RP-18 endcapped 100–2 mm, Sigma 152006) and introduced into a JetStream source (Agilent) for triple quadrupole analysis. Data were analyzed and quantified using MassHunter software (Agilent)^{39 (link),40 (link)}.