Hair cortisol (ancillary study)

Hair cortisol (ancillary study)

LASA filenames:

Contact: Natasja van Schoor


Cortisol, commonly known as “the stress hormone” is often used to assess the stress level in humans. Until a few years ago, cortisol has solely been analyzed from blood serum, saliva, or urine. These analyses offer the possibility to explore the dynamics and the concentration of acutely (serum, saliva) or short-term (urine) circulating cortisol concentrations. Unfortunately, the routine methods of measuring cortisol have drawbacks as they show considerable intra- and interindividual differences which are due to cortisol’s circadian rhythm as well as its pulsatile secretion (1), daily variation, and reactivity to acute transient stress, such as nervousness (2). These techniques further call for invasive or frequent sampling and they are especially prone to measurement error and sloppiness as the samples are often collected by the participants themselves, without supervision (4).

For a few years now, a new and very different method to measure cortisol exposure in humans has been developed; the extraction of cortisol from human hair, with first evidence provided in 2004 (5). Since then, several research groups have been focusing on this technique with some of its numerous advantages being the noninvasiveness, the standardized sampling, and, maybe most intriguing, the possibility to use hair as a retrospective biomarker of cortisol exposure. As hair grows approximately one centimeter per month (5), hair analysis offers the possibility to show the average long-term activity of the HPA axis, and to compare several hair segments/months with each other. Hair cortisol most probably reflects the amount of free, unbound cortisol (6).

Measurements in LASA

Hair collection, preparation, and cortisol measurement

For each respondent, around 150 strands of hair were collected from the posterior vertex of the scalp and were cut off as close to the scalp as possible. The hair was taped to a piece of paper and the scalp end was marked. The samples were stored in an envelope at room temperature until analysis.

The proximal 3 cm of hair, reflecting roughly the 3 months before hair sample collection, were used for the measurement of cortisol. A minimum of 15 mg of hair was weighed and cut in small pieces in a glass vial. 1 mL of methanol was added and the vial was sealed and incubated overnight for 16 h at 52 °C while gently shaking. After incubation, the methanol was removed, put into disposable glass tubes and was evaporated under constant stream of nitrogen. The samples were dissolved in 250 μL phosphate buffered saline (pH 8.0) and samples were vortexed for 1 min. Before analysis, the samples were vortexed again for 30 s. Cortisol levels were measured using a commercially available ELISA Kit for Salivary Cortisol (DRG Instruments GmbH, Marburg, Germany) as per manufacturer’s directions with the reagents provided. Cross reactivity of other steroids with the kit’s antibodies was reported as follows: Corticosterone (29.00%), Cortisone (3.00%), 11-Deoxycortisol (<1.00%), 17-OH Progesterone (<0.50%), other hormones (<0.10%). Intra-assay variation was below 5% and the inter-assay variation below 8% as stated by the manufacturer.


A self-developed questionnaire regarding hair characteristics and hair treatment was used. Participants had to provide information on their natural hair colour (black, brown, blond, red, grey), whether the hair was treated in the last six months (dyed, bleached, permed) and if yes, which treatment it had undergone. The third question asked was when this treatment had been applied the last time (< 1 month ago, 1-2 months ago, 3-4 months ago, and > 4 months ago). The fourth question was about the frequency of hair washing (< 1 times a week, 1-2 times a week, 3-4 times a week, > 4 times a week). The fifth question was whether on the day of hair assessment, a hair product has been used, and if yes, which one (mousse, gel, wax, spray, other, more than one). The next question was whether in the last three months, corticosteroids had been used, and if yes, which sort (pills, inhalation, ointment, nose spray, other).

This information is required as bleaching, dyeing and hair washing frequency have inconsistently shown to be associated with altered hair cortisol levels and may need to be adjusted for. Corticosteroids do alter hair cortisol levels and use of them are often considered an exclusion criterion.

Variable information

LASAGEPOSA883_varinfo (pdf)

Availability of data per wave
Data on hair cortisol is collected during the baseline study of the European Project on OSteoArthritis (EPOSA) (2010-2011), which is a side study of LASA ( All 574 Dutch EPOSA participants at baseline were asked to participate in a study on hair cortisol. In total, data on hair cortisol was collected for 333 respondents.

Previous use in LASA

The hair cortisol data have been used in one study until now:


  1. Lightman, S.L., Wiles, C.C., Atkinson, H.C., Henley, D.E., Russell, G.M., Leendertz, J.A., McKenna, M.A., Spiga, F., Wood, S.A., Conway-Campbell, B.L., 2008. The significance of glucocorticoid pulsatility. Eur. J. Pharmacol. 583, 255-262.
  2. Hellhammer, J., Fries, E., Schweisthal, O.W., Schlotz, W., Stone, A.A., Hagemann, D., 2007. Several daily measurements are necessary to reliably assess the cortisol rise after awakening: state- and trait components. Psychoneuroendocrinology 32, 80-86.
  3. Lightman, S.L., Wiles, C.C., Atkinson, H.C., Henley, D.E., Russell, G.M., Leendertz, J.A., McKenna, M.A., Spiga, F., Wood, S.A., Conway-Campbell, B.L., 2008. The significance of glucocorticoid pulsatility. Eur. J. Pharmacol. 583, 255-262.
  4. Kudielka, B.M., Broderick, J.E., Kirschbaum, C., 2003. Compliance with saliva sampling protocols: electronic monitoring reveals invalid cortisol daytime profiles in noncompliant subjects. Psychosom. Med. 65, 313-319.
  5. Raul, J.S., Cirimele, V., Ludes, B., Kintz, P., 2004. Detection of physiological concentrations of cortisol and cortisone in human hair. Clin. Biochem. 37, 1105-1111.
  6. Dettenborn, L., Tietze, A., Kirschbaum, C., Stalder, T., 2012. The assessment of cortisol in human hair: associations with sociodemographic variables and potential confounders. Stress 15, 578-588.
  7. Gow, R., Thomson, S., Rieder, M., Van Uum, S., Koren, G., 2010. An assessment of cortisol analysis in hair and its clinical applications. Forensic Sci. Int. 196, 32-37.
  8. Manenschijn, L., Koper, J.W., Lamberts, S.W.J., van Rossum, E.F.C., 2011a. Evaluation of a method to measure long term cortisol levels. Steroids 76, 1032-1036.

Date of last update: April 7, 2020