Establishment of reference intervals for sex hormone binding globulin, LC-MS/MS derived free androgen index and calculated free testosterone SN Duignan, MM MacMahon, KJ Mulready, JJ Brady, MC Fitzgibbon, MR Cullen Department of Clinical Biochemistry & Diagnostic Endocrinology, Mater Misericordiae University Hospital, Dublin Introduction We have recently introduced a liquid chromatography–tandem e mass spectrometry (LC-MS/MS) total testosterone (T) assay into into routine clinical practice. This method is highly sensitive and specifi and is superior to immunoassay for measuring T Analysis of sex hormone binding globulin (SHBG), and either determining the free androgen index (FAI) in females or the calculated free T (cFT) in males is used to support the clinical interpretation of equivalent T levels Our aim was to derive gender-specific SHBG, LC-MS/MS FAI and cFT reference intervals (RI) M Reference interval distribution plots Females Males Testosterone Method Seventy five females and 38 males with an age range spanning 20 to 61 years were recruited. Exclusion criteria included hormonal/steroid therapy (including OCP), drugs (including thyroxine and anti-convulsants), albumin <35 g/L, and known Polycystic Ovary Syndrome T was measured by an automated reverse phase/ion-exchange extraction procedure and analyzed by chromatography using an Acquity UPLC linked to a Waters Xevo tandem mass spectrometer. SHBG was measured using immunoassay (IA) on an Abbott Architect i2000sr The FAI was used to determine the free T in females1. The Vermuelen equation was used to determine the cFT in males2 The female FAI RI was validated by: Comparing T and FAI determined by IA at MMUH and by LC-MS/MS at a referral lab over a two year period, n=186 Auditing the clinical presentation and imaging, where available, of all FAI’s since the introduction of the new LC-MS/MS assay at MMUH, n=66 Relationship between FAI and CFT Validation of LC-MS/MS Derived FAI Results The SHBG RI’s for females and males were 20.9–226.5 nmol/L and 5.9–68.5 nmol/L respectively. The RI’s for the female FAI and for the male cFT were 0.29–5.26% and 0.22-0.93 nmol/L respectively The FAI correlated well with cFT in females (r=0.945). In males, the FAI was shown to correlate poorly with cFT (r=0.184) and over estimate cFT at low SHBG concentrations Our new LC-MS/MS FAI RI is lower than the previous upper limit of the MMUH (8.5%) which was determined using IA. We validated our new LC-MS/MS FAI RI by confirming an observed positive bias over a 2 year period for IA compared to LC- MS/MS values. In addition, we found that by applying the new LC-MS/MS FAI, the clinical sensitivity increases without compromising the clinical specificity Method Comparison Clinical Performance Conclusion We have derived gender specific SHBG RI’s and also LC-MS/MS derived female FAI and male cFT RI’s in a healthy population. The female SHBG RI differed significantly from the manufacture’s quoted interval (19.8-155.2 nmol/L). Our new RI may benefit from pre- and post-menopausal partitioning Our LC-MS/MS FAI and cFT will improve the diagnostic efficiency of interpreting equivalent T levels when introduced into routine clinical practice. References GD Carter et al. Investigation of hirutism: testosterone is not enough. Ann Clin Biochem 1983; 262-263 Vermeulen et al. A critical evaluation of simple methods for the estimations of free testosterone in serum. J Clin Endocrinol 1999; 84 3666-3672