Leveraging mass spectrometry for accurate diagnosis of pediatric endocrine disorders

Pediatric endocrine disorders are conditions characterized by disruptions in hormonal regulation.¹ Examples include congenital adrenal hyperplasia (CAH), abnormalities in pubertal timing, differences in sex development, and specifically in adolescents, polycystic ovary syndrome (PCOS). Accurate diagnosis and effective treatment require careful biochemical monitoring. However, a major challenge in managing these disorders is the complexity of steroid hormone analysis.² Many of these disease-driving steroid hormones are present in very low concentrations and have similar molecular structures, making them difficult to detect and measure.^3,4 As a result, hormone testing methods must offer both high specificity and sensitivity.

Medical laboratories commonly use immunoassays to manage pediatric endocrine disorders due to their scalability and compatibility with high-throughput analyzers. While convenient, immunoassays may struggle to measure hormones at low concentrations accurately and can be subject to interference from other biological compounds.³ Liquid chromatography-tandem mass spectrometry (LC-MS/MS), by contrast, offers high sensitivity and specificity, enabling the detection of low-abundance molecules.⁴ LC-MS/MS is becoming increasingly available as a tool for improving the diagnosis and management of pediatric endocrine disorders.⁵

Researchers and healthcare institutions rely on LC-MS/MS for a broad range of diagnostic applications in pediatric endocrinology.⁶ This includes accurately quantifying steroid hormones, detecting rare metabolic disorders, and monitoring hormone replacement therapies. Its ability to deliver precise results makes LC-MS/MS valuable for diagnosing conditions where hormone levels are low, fluctuate rapidly, or change during a patient’s lifetime.

CAH encompasses a group of inherited disorders characterized by impaired adrenal steroidogenesis, with 21-hydroxylase deficiency (21OHD) accounting for the majority of cases. Early referral to experienced multidisciplinary teams alongside comprehensive biochemical testing ensures timely and appropriate intervention. Accurate steroid hormone analysis is needed for precise diagnosis.⁷

Initial screening and diagnosis typically involve immunoassay-based measurement of 17-hydroxyprogesterone (17OHP) hormone levels.⁸ Increasing evidence supports LC-MS/MS for quantifying 17OHP and related adrenal steroids, providing improved analytical precision and helping to confirm immunoassay results.⁹ This approach enhances diagnostic accuracy, informs therapeutic decision-making, and supports more precise monitoring of treatment efficacy. 

Close biochemical surveillance in CAH, enabled by mass spectrometry, can optimize glucocorticoid dosing, minimize risks of overtreatment, and facilitate shared decision-making between healthcare providers and families.⁷

LC-MS/MS methodologies are increasingly integrated into clinical practice for CAH management, offering reduced false-positive rates and higher diagnostic confidence. The Endocrine Society and the Society for Endocrinology UK recommends LC-MS/MS over immunoassays due to its superior specificity, sensitivity, and performance in measuring key adrenal steroids.^8,10

Vitamin D deficiency can significantly impact bone density and increase the risk of serious conditions such as osteoporosis.¹¹ A major challenge in diagnosing deficiency and monitoring treatment response is the accurate measurement of vitamin D metabolites, which has been complicated by cross-reactivity issues in traditional assays.^12,13 Studies show that automated LC-MS/MS enables precise quantification of 25-hydroxyvitamin D [25(OH)D], the primary biomarker for assessing vitamin D status, overcoming long-standing difficulties caused by assay variability and lack of standardization.⁵ By minimizing cross-reactivity and reducing overestimation compared to immunoassays, LC-MS/MS is particularly valuable for vulnerable populations, including infants under one year of age.

Clinical laboratories are adopting LC-MS/MS because it offers more reliable diagnoses and management of vitamin D deficiency, especially needed due to inaccurate detection from cross-reactivity of vitamin D metabolites.

PCOS is a common endocrine disorder affecting up to 13% of reproductive‐aged women that cause irregular menstrual cycles and hyperandrogenism, among other symptoms. The heterogeneity of symptoms, the multisystem nature of PCOS, and similar clinical presentation with other conditions make it challenging to achieve an accurate diagnosis.¹⁴

It is becoming increasingly evident that LC-MS/MS has high sensitivity and specificity for diagnosing hyperandrogenism, offering more accurate quantification of PCOS-related androgens such as total testosterone, free testosterone, androstenedione, and dehydroepiandrosterone sulfate (DHEAS).^15,16 This helps reduce diagnostic variability, improves the reliability of PCOS diagnosis, and informs more precise clinical management.

While mass spectrometry has been traditionally reserved for research purposes or in specialized clinical labs, automation is now increasing access for conventional labs, especially critical for the field of pediatric endocrinology disorders. Automation of this highly complex process and technology reduces labor-intensive manual workflows and can be expanded to more urgent and routine testing for higher throughput.^17,18 Additionally, automation is helping laboratories more easily incorporate mass spectrometry into their existing routine laboratory and laboratory information management systems. 

However, before the widespread adoption of automated mass spectrometry, there was a need for standardizing processes. Today, variability across labs for identifying and quantifying steroid hormones leads to inconsistent results. Mass spectrometry, while uniquely capable of highly sensitive and specific detection, is susceptible to potential sources of error and inaccuracy.¹⁹ Without the standardization of reference material and procedures, results can vary and make clinical diagnoses unreliable. 

Automated mass spectrometry platforms are becoming more accessible in clinical settings. Laboratory leaders can leverage this technology to ensure more accurate hormone quantification, validate immunoassay results, and implement robust testing protocols. These efforts help physicians make more informed diagnostic and treatment decisions for pediatric endocrine disorders, directly impacting patient care.