April 11, 2024 | As the body’s stress hormone, cortisol influences several diseases, from chronic illnesses, such as diabetes and cancer; to mental illnesses, such as depression; to neurological diseases, such as Alzheimer’s disease. Researchers at Aarhus University have discovered a breakthrough method to measure cortisol in an efficient, faster way. This new analysis test provides quicker and more accurate diagnoses, allowing clinicians to help their patients in a timelier manner.
Greater Efficiency and Faster Results
“It was a side project, actually,” says Mathias Flensted Poulsen, one of the research team members, about how the study began. “We wanted an in-house bioassay to test something completely else.”
The initial in-house bioassay was a cell-based glucocorticoid reporter assay. After it was validated with dexamethasone, the team wondered if serum from blood samples could be tested and how it would react. The result was that it “worked well.”
Despite the humble beginnings of the project, it uncovered a promising lead in diagnoses in clinical practice. Compared to the current method of measuring cortisol, the new method is less tedious and time-consuming (Analytical Chemistry, DOI: 10.1021/acs.analchem.3c04435). The current method is described as laborious, unreliable, and inconvenient and requires purifying the sample via chromatography purification dialysis or ultracentrifugation before analyzing for bioactive cortisol. The issue with purification is that a part of the sample is lost with each step. An experienced researcher can ensure purification goes smoothly, but it is nonetheless susceptible to human error, especially when a large sample size is involved.
The new method does not require a long purification process. Instead, a serum sample is applied to the bioassay cells. Unlike immunoassays and mass spectrometry, the bioassay cells do not directly quantify the exact concentration of cortisol. Rather, the serum influences the cells to sense natural and unnatural glucocorticoids, thus assessing bioactivity of the total glucocorticoid load that encompasses both cortisol and synthetic glucocorticoids from medication in patients taking exogenous glucocorticoids, according to Poulsen. Specifically, the serum introduces a protein called luciferase, and when luciferin, the substrate for luciferase, is added, it causes an enzymatic reaction that produces light—much like how a firefly emits light. This light produced by the cells is then measured via a standard plate reader, compared to a standard curve of known cortisol concentrations, and translated into a cortisol concentration.
In a clinical setting, the results obtained from the bioassay closely resemble the way immunoassays obtain results. The workflow and analysis procedures are quite similar between the two. However, the bioassay provides a more physiological, relevant estimate of what is in the blood sample and what activates the glucocorticoid pathway instead of giving an exact concentration of cortisol levels.
“The unexpected part of this was actually how smoothly it was,” comments Poulsen on the process and discovery. “I have never experienced that before—that when you start something up, and it kind of just works.”
Bringing the Test to Clinics
Ultimately, the team wants to bring the test to clinics and give physicians an easier method of measuring cortisol, particularly with glucocorticoid treatments. “We hope that… it can give some predictive value and more personalized regimes of treatment with glucocorticoids, that the free glucocorticoid measurement can predict if people are on the right dosage,” says Poulsen. There is also the potential of using the test to predict patients who have lower or higher glucocorticoid responses, which would help with adjusting dosages for patients.
But there are, of course, obstacles to overcome. As of right now, clinics do not use cell-based assays, and there could be optimization problems. However, Poulsen is hopeful that the test “should be relatively easy” to distribute to clinics once more studies are conducted, which the team is already in the process of planning, and more information is acquired.
However, Poulsen warns that while the results of the study show great promise, the method is still in the preliminary stages. Nonetheless, as the team plans for future projects, clinicians and physicians can keep an eye out for new developments.
“For now, it just shows that it measures bioactive glucocorticoid and is pretty good at it compared to the immunoassays and the mass spectrometry,” says Poulsen.