CORTISOL PORTABLE BIOSENSORS

HERE IS SOME RESEARCH AIMED AT ALLOWING AN INDIVIDUAL TO SEE CORTISOL LEVELS WITHOUT SENDING THE SALIVA CORTISOL SAMPLE OFF TO A LAB.

Anal Chem. 2008 Jul 26. [Epub ahead of print]
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Detection of Cortisol in Saliva with a Flow-Filtered, Portable Surface Plasmon Resonance Biosensor System.Stevens RC, Soelberg SD, Near S, Furlong CE.clem@u.washington.edu.Saliva provides a useful and noninvasive alternative to blood for many biomedical diagnostic assays. The level of the hormone cortisol in blood and saliva is related to the level of stress. We present here the development of a portable surface plasmon resonance (SPR) biosensor system for detection of cortisol in saliva. Cortisol-specific monoclonal antibodies were used to develop a competition assay with a six-channel portable SPR biosensor designed in our laboratory. The detection limit of cortisol in laboratory buffers was 0.36 ng/mL (1.0 nM). An in-line filter based on diffusion through a hollow fiber hydrophilic membrane served to separate small molecules from the complex macromolecular matrix of saliva prior to introduction to the sensor surface. The filtering flow cell provided in-line separation of small molecules from salivary mucins and other large molecules with only a 29% reduction of signal compared with direct flow of the same concentration of analyte over the sensor surface. A standard curve for detection of cortisol in saliva was generated with a detection limit of 1.0 ng/mL (3.6 nM), sufficiently sensitive for clinical use. The system will also be useful for a wide range of applications where small molecular weight analytes are found in complex matrixes.

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ASAP Anal. Chem., ASAP Article, 10.1021/ac800892h Web Release Date: July 26, 2008 Copyright Sciences, University of Washington,


The steroid hormone cortisol is required for the regulation of blood pressure, cardiovascular function, and many metabolic activities. The body regulates cortisol levels by controlling cortisol secretion and production based on levels in the bloodstream.1 An excess of cortisol leads to Cushing’s syndrome with symptoms that include obesity, fatigue, and skin and bone fragility.2 A deficiency in cortisol leads to Addison’s disease with symptoms including weight loss, fatigue, and darkening of skin folds and scars.3 Normal serum levels of cortisol vary on a diurnal cycle from 30 to 140 ng/mL (100−500 nM), peaking in early morning.4 Cortisol levels increase in response to physical stresses such as injury and illness as well as psychological stresses including depression and fatigue.5 Measuring serum cortisol levels has become an important diagnostic indicator of overall stress as well as disease state of patients.
Analysis of cortisol in saliva is a useful alternative to the standard method of measuring serum cortisol levels for several reasons. First, serum cortisol measurements include the cortisol bound to transport proteins and albumin while the fraction of free cortisol is thought to be more biologically active.6 Also, levels of salivary cortisol have been shown to correlate well with levels of unbound serum cortisol;7, 8 however, overall levels of cortisol in saliva are up to 100-fold lower than in serum.9 Morning salivary cortisol levels range from 1 to 8 ng/mL (3.5−27 nM) in healthy adults.9 Finally, methods used for collecting blood may create stress and result in misleading cortisol values.10, 11 It is also difficult to collect serum samples over the course of a whole day, while collecting saliva is easy and the samples remain useful for analysis for up to one week following collection.12 Methods for determining salivary cortisol provide an opportunity to increase ease and accuracy in detecting disease or stress level.
Several commercial kits use competitive enzyme-linked immunosorbant assay (cELISA) technology to measure cortisol in saliva, including kits available from Salimetrics (State College, PA) and Orion Diagnostica (Espoo, Finland). The cELISA methods provide accurate and reproducible results but require several hours for analysis. Other immunoassays have been used to measure salivary cortisol levels including time-resolved fluoroimmunoassay,13 rapid quantitative immunodetection using a lateral flow assay,14 and luminescence immunoassays.15, 16 These immunoassays also take considerable time or lack quantifiable results. One protocol for detecting salivary cortisol uses liquid chromatography−tandem mass spectrometry but requires a slow cortisol extraction step.17 A near real-time method for detection of cortisol in saliva with little sample preparation has not yet been reported.
Surface plasmon resonance (SPR) detection of cortisol provides a method for rapid measurement of cortisol in saliva. SPR technology measures the changes in refractive index (RI) near the sensor surface in response to molecular interactions. Specific receptor molecules can be immobilized onto the gold sensor surface for detecting target analytes in solution. Binding of targets to the receptors results in measurable changes in RI. Binding of large target analytes such as proteins, viruses, spores, and microbes to specific antibodies immobilized on the gold surface is easily monitored in near real time. Due to the significant change in RI occurring when these analytes are bound, labels or amplifying reagents are unnecessary for many SPR applications. A general review of SPR can be found in a book edited by Davies.18 Small organic molecules such as cortisol require different methods for detection since direct binding of small targets to receptor molecules does not result in large changes in RI. As a result, competition or displacement assays are used to detect small analytes at clinically relevant levels. Competition assays are based on the inhibition of antibody binding to immobilized target by free analyte in solution. SPR competition assays have been used to successfully detect small molecules including the herbicide atrazine,19 the pesticides fenitrothion,20 DDT, chlorpyrifos, and carbaryl,21 the pesticide metabolite paraoxon,22 the antiepileptic drug phenytoin,23 the hormones thyroxine24 and estradiol,25 and the shellfish toxin domoic acid.26, 27
Here, we report the detection of cortisol in saliva using a portable SPR biosensor and competition-based assays combined with a specially designed flow cell that reduces nonspecific binding by retaining large molecules in the sample but delivering small molecular weight analytes to the sensor surface.


CONCLUSIONS
We report here a rapid assay for measuring cortisol levels in saliva. The small, portable SPR biosensor used for measurements of cortisol will allow for analysis in the field as well as in emergency rooms and provides results in less than 10 min. Levels of cortisol in blood and saliva can be measured and used as an indicator of stress. While measurements of cortisol in saliva are as much as 100× lower than in blood, it is thought that salivary cortisol and free cortisol in blood correlate more closely with stress than does total blood cortisol.6 Also, collecting blood may induce stress and alter the measurement while saliva collection is stress-free. The collection of saliva does not require trained medical skills and can, therefore, simplify collection of samples at numerous time points.
The in-line filtering flow cell described here will be useful for detecting many other small molecular weight analytes in complex matrixes. Detection of cortisol in a complex matrix such as saliva has been simplified by the addition of the external in-line filtering flow cell that allows for diffusion and bulk flow of small molecular weight analytes into the sensor feed stream while excluding particulates and larger organic molecules that can cause an increase in background signal. The external in-line filtering flow cell can be easily adapted for use in other SPR devices as well as many other diagnostic biosensors with upstream processing capabilities



2: Psychol Assess. 2007 Mar;19(1):44-57.
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Clinical assessment applications of ambulatory biosensors.Haynes SN, Yoshioka DT.Department of Psychology, University of Hawaii at Manoa, Honolulu, HI 96822, USA. sneil@hawaii.eduAmbulatory biosensor assessment includes a diverse set of rapidly developing and increasingly technologically sophisticated strategies to acquire minimally disruptive measures of physiological and motor variables of persons in their natural environments. Numerous studies have measured cardiovascular variables, physical activity, and biochemicals such as cortisol in psychopathology and treatment research. The physiological concomitants of many behavior and medical disorders and the benefits of a multimethod assessment strategy provide strong rationales for clinical applications of ambulatory biosensor measurement. A number of psychometric dimensions of evaluation are important in clinical applications of biosensor measurement, including accuracy and validity, reliability and consistency, clinical utility, incremental validity and utility, sensitivity to change, generalizability, cost benefits, and the conditional nature of dimensions of biomeasure evaluation. The authors review ambulatory biosensor methods and make recommendations for use of the technology. ((c) 2007 APA, all rights reserved).Publication Types:
Review