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Updated: Apr 15, 2026
Collecting Hair Samples for Hair Cortisol Analysis in African Americans
Published on: June 10, 2018
Methodological Considerations for Hair Cortisol Measurements in Children
Radomir Slominski1, Cynthia R Rovnaghi, Kanwaljeet J S Anand
1*Pain Neurobiology Laboratory, Department of Pediatrics, Neuroscience Institute, University of Tennessee Health Science Center; and †Division of Pediatric Critical Care Medicine, Department of Pediatrics, University of Tennessee Health Science Center and Le Bonheur Children's Hospital, Memphis, TN.
Optimizing hair cortisol analysis is crucial for accurate chronic stress assessment in children. Modified extraction methods, including sequential incubations, improve cortisol yield and reliability for diverse populations.
Area of Science:
- Biochemistry
- Endocrinology
- Analytical Chemistry
Background:
- Hair cortisol analysis is increasingly used to assess chronic stress in pediatric populations.
- Current methodologies may limit the accuracy of reference ranges across diverse groups.
- Standardized and optimized methods are needed for reliable hair cortisol quantification.
Purpose of the Study:
- To compare different hair processing techniques (milled vs. finely cut) for cortisol extraction.
- To develop and evaluate an optimized sequential extraction process for hair cortisol and protein.
- To compare the performance of liquid chromatography-mass spectrometry (LC-MS) and enzyme-linked immunosorbent assays (ELISAs) for hair cortisol measurement.
Main Methods:
- Comparison of finely cut versus milled hair processing methods.
- Development of a 4-step sequential extraction protocol using methanol and acetone.
- Comparative analysis of cortisol and protein yields using spectrophotometry and ELISA.
- Validation of ELISA results against LC-MS for hair cortisol quantification.
Main Results:
- Finely cut hair processing yielded higher cortisol and protein compared to milled methods.
- Sequential extraction significantly increased cortisol and protein recovery, with diminishing returns after the third incubation.
- ELISA demonstrated higher sensitivity in detecting hair cortisol levels compared to LC-MS.
- LC-MS identified cortisone as a significant steroid component alongside cortisol.
Conclusions:
- Finely cutting hair and employing sequential extractions (at least 3 incubations) enhance cortisol yield.
- Optimized extraction protocols are essential for establishing accurate hair cortisol reference ranges.
- ELISA offers superior sensitivity for hair cortisol measurement compared to LC-MS, aiding in stress assessment.
