Accuracy of Testosterone and Androstenedione in PCOS Diagnosis

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Accuracy of Testosterone and Androstenedione in PCOS diagnosis

Hyperandrogenism by Liquid Chromatography Tandem Mass Spectrometry in PCOS: Focus on Testosterone and Androstenedione

A single centre prospective study of female patients was conducted to evaluate the impact of using LC-MS/MS (liquid chromatography in tandem with mass spectrometry) for diagnosis of PCOS, while also determining which androgen can identify PCOS women from controls most accurately.

Although LC-MS/MS is not new technology, it has limited use in clinical settings due to the complexity associated with preparing samples and unambiguous cut-off values. A recent commercial product, which was produced to simplify this process, was used in this study.

To properly test this, 117 women suspected of PCOS who met the inclusion criteria, participated in the study along with 73 women, without PCOS, as controls. Classification of PCOS or non-PCOS was initially based on testosterone, free Testosterone and Androstenedione levels by immunoassays. These androgens was then measured by LC-MS/MS to evaluate its potential impact on diagnosis rates.

Initial results showed that the concentration of androgens tended to be lower, when measured by LC-MS/MS, compared to immunoassay, however this was only statistically significant for Androstenedione. Overall PCOS was diagnosed in 90 of the 117 women by immunoassay, while LC-MS/MS identified 93 of the 117 women as having PCOS.

Interestingly LC-MS/MS analysis identified significantly more women with the classical type of PCOS (oligoamenorrhea, hyperandrogenism) compared to immunoassays (77.1 vs 55.6%). Hyperandrogenism, either elevated testosterone, free Testosterone or Androstenedione, by LC-MS/MS was found in 89.2% of women with PCOS. Of the 3 androgens, high free Testosterone was the most prevalent (73.1%), followed by Androstenedione (64.5%).

The sensitivity, specificity and AUC (Area under the ROC curve) listed below, with specific cut-offs, shows the predictive performance of the 3 androgens along with the Free Androgen Index, in differentiating PCOS.

Cut-offSensitivity (%)Specificity (%)AUC
Testosterone0.24 ng/mL87750.899
free Testosterone0.34 ng/dL91890.954
Free Androgen Index1.6790830.951
Androstenedione1.16 ng/mL91880.957

Further analysis found only free Testosterone (and not testosterone or Androstenedione) correlated with clinical and metabolic parameters of PCOS, specifically BMI, HOMA-Index, HbA1c and SHBG.

Overall the findings support the use of Androstenedione to identify hyperandrogenism in women with PCOS, with Androstenedione as accurate as free Testosterone in differentiating PCOS.


According to this study, which analysed 93 women with PCOS, elevated levels of free Testosterone was significantly more prevalent than elevated Total Testosterone (73.1% vs 61.3%), in women with PCOS. Similarly, the predictive power for PCOS of free Testosterone is higher than Total Testosterone (0.954 vs 0.899).


  1. PCOS only defined by Rotterdam criteria
  2. Lean cohort of women only

Similar studies

Handelsman D J, et al. (2017). Performance of mass spectrometry steroid profiling for diagnosis of polycystic ovary syndrome.

Tosi F, et al. (2016). Implications of Androgen Assay Accuracy in the Phenotyping of Women With Polycystic Ovary Syndrome.

Pasquali R, et al. (2016). Defining Hyperandrogenism in Women With Polycystic Ovary Syndrome: A Challenging Perspective.

Fanelli F, et al. (2011). Serum steroid profiling by isotopic dilution-liquid chromatography–mass spectrometry: Comparison with current immunoassays and reference intervals in healthy adults.


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