Methods in the study of marine mammal stress: Measuring binding affinity of corticosteroid binding globulin

Delehanty, Brendan; Bossart, Gregory D.; Champagne, Cory D.; Crocker, Daniel E.; Fair, Patricia A.; Haulena, Martin; Houser, Dorian S.; Richardson, Evan S.; Lunn, Nicholas J.; Boonstra, Rudy

doi:10.1111/mms.12612

Cited by 4 publications

(8 citation statements)

References 30 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…However, if too much [ 3 H]-CORT is added, measurement error increases. We identified the optimal saturation concentration by examining the saturation binding curve that was used to determine the equilibrium dissociation constant ( K d ) for the species of interest (see Delehanty et al ., 2015 and Delehanty et al ., 2019 for details on performing saturation binding assays). The saturation binding curve is produced by incubating plasma with an increasing amount of [ 3 H]-CORT.…”

Section: Methodsmentioning

confidence: 98%

“… Saturation binding curve at 4°C for Antarctic fur seal (data from Delehanty et al ., 2019 ). Specific binding (by CBG) is the difference between total binding and nonspecific binding.…”

Section: Methodsmentioning

confidence: 99%

“…Fig. 1 is a saturation binding curve for the Antarctic fur seal (data from Delehanty et al ., 2019 ). Note that the x-axis is the free hormone concentration in the plasma mixture at equilibrium, not the total CORT concentration added to the mixture: a portion of the CORT added to the plasma solution is bound to the CBG.…”

Section: Methodsmentioning

confidence: 99%

“…As explained in the Methods and Supporting File 1, the best way to determine an optimal saturating concentration of [ 3 H]-CORT is to incubate diluted plasma with increasing concentrations of [ 3 H]-CORT to determine at what point specific binding is maximized. Fortunately, this is exactly what is done in a saturation binding curve that is used to determine the binding affinity (K d ) of CBG (see Delehanty et al ., 2015 ; Delehanty et al ., 2019 for that assay, and see below to see how K d fits into the calculation of free hormone. We explain in the Methods section how a saturation binding curve can be used to determine the saturating [ 3 H]-CORT concentration.…”

Section: Basic Principles Of the Assaysmentioning

confidence: 99%

“…Once each of these values is determined, the free CORT concentration can be calculated using a formula based on the laws of mass action ( Barsano and Baumann, 1989 ). We have previously published comprehensive accounts of the methods to calculate the binding affinity (represented by the equilibrium dissociation constant, K d ) of CBG for a wide range of species (terrestrial mammals in Delehanty et al ., 2015 and marine mammals in Delehanty et al ., 2019 ).…”

Section: Introductionmentioning

confidence: 99%

See 4 more Smart Citations

Measurement of free glucocorticoids: quantifying corticosteroid binding capacity and its variation within and among mammal and bird species

Delehanty

Bossart

Champagne

et al. 2020

Conservation Physiology

Self Cite

View full text Add to dashboard Cite

Plasma glucocorticoid (CORT) levels are one measure of stress in wildlife and give us insight into natural processes relevant to conservation issues. Many studies use total CORT concentrations to draw conclusions about animals’ stress state and response to their environment. However, the blood of tetrapods contains corticosteroid-binding globulin (CBG), which strongly binds most circulating CORT. Only free CORT (CORT not bound by CBG) leaves the circulation and exerts biological effects on CORT-sensitive tissues. Measuring free CORT concentrations provides insight to an animal’s stress response that cannot be revealed by simply measuring total CORT. To calculate free CORT concentrations in plasma or serum samples, one needs three measurements: the binding affinity of CBG for CORT (which varies by species), the total CORT concentration in the sample and the maximum corticosteroid binding capacity (MCBC) of CBG in the sample. Here, we detail the measurement of CBG binding capacity. We compare and contrast the three main methods to measure MCBC: charcoal, cell harvester and dialysis. Each is defined by the means by which free and bound CORT are separated. We weigh the relative merits and challenges of each. We conclude that sample volume, species and taxon binding specificity, and availability of equipment are the primary considerations in selecting the appropriate separation method. For most mammals, the charcoal method is recommended. For birds, the harvester method has critical advantages over the charcoal method. The dialysis method is widely regarded as the gold standard and has lower equipment costs but is more time-intensive and costly in terms of radioactive isotope needed and is less suited to processing large numbers of samples. The binding capacity of CBG varies tremendously within and among the bird and marine mammal species studied, and we discuss the implication of this variation for understanding the role of stress in wildlife.

show abstract

Section: Methodsmentioning

confidence: 98%

“… Saturation binding curve at 4°C for Antarctic fur seal (data from Delehanty et al ., 2019 ). Specific binding (by CBG) is the difference between total binding and nonspecific binding.…”

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Basic Principles Of the Assaysmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 3 more Smart Citations

Measurement of free glucocorticoids: quantifying corticosteroid binding capacity and its variation within and among mammal and bird species

Delehanty

Bossart

Champagne

et al. 2020

Conservation Physiology

Self Cite

View full text Add to dashboard Cite

show abstract

Immunology

De Guise,

Levin,

Romano

et al. 2024

The Physiology of Dolphins

View full text Add to dashboard Cite

Investigation of the Link between Per- and Polyfluoroalkyl Substances and Stress Biomarkers in Bottlenose Dolphins (Tursiops truncatus)

Bennett,

Aung,

Boonstra

et al. 2024

Environ. Sci. Technol.

Self Cite

View full text Add to dashboard Cite

Bottlenose dolphins (Tursiops truncatus) are keystone and sentinel species in the world’s oceans. We studied correlations between per- and polyfluoroalkyl substances (PFAS) and their stress axis. We investigated associations between plasma biomarkers of 12 different PFAS variants and three cortisol pools (total, bound, and free) in wild T. truncatus from estuarine waters of Charleston, South Carolina (n = 115) and Indian River Lagoon, Florida (n = 178) from 2003 to 2006, 2010–2013, and 2015. All PFAS and total cortisol levels for these dolphins were previously reported; bound cortisol levels and free cortisol calculations have not been previously reported. We tested null hypotheses that levels of each PFAS were not correlated with those of each cortisol pool. Free cortisol levels were lower when PFOS, PFOA, and PFHxS biomarker levels were higher, but free cortisol levels were higher when PFTriA was higher. Bound cortisol levels were higher when there were higher PFDA, PFDoDA, PFDS, PFTeA, and PFUnDA biomarkers. Total cortisol was higher when PFOA was lower, but total cortisol was higher when PFDA, PFDoDA, PFTeA, and PFTriA were higher. Additional analyses indicated sex and age trends, as well as heterogeneity of effects from the covariates carbon chain length and PFAS class. Although this is a cross-sectional observational study and, therefore, could reflect cortisol impacts on PFAS toxicokinetics, these correlations are suggestive that PFAS impacts the stress axis in T. truncatus. However, if PFAS do impact the stress axis of dolphins, it is specific to the chemical structure, and could affect the individual pools of cortisol differently. It is critical to conduct long-term studies on these dolphins and to compare them to populations that have no or little expose to PFAS.

show abstract

Methods in the study of marine mammal stress: Measuring binding affinity of corticosteroid binding globulin

Cited by 4 publications

References 30 publications

Measurement of free glucocorticoids: quantifying corticosteroid binding capacity and its variation within and among mammal and bird species

Measurement of free glucocorticoids: quantifying corticosteroid binding capacity and its variation within and among mammal and bird species

Immunology

Investigation of the Link between Per- and Polyfluoroalkyl Substances and Stress Biomarkers in Bottlenose Dolphins (Tursiops truncatus)

Contact Info

Product

Resources

About