Plasma adrenocorticotropic hormone (ACTH) concentrations in ponies measured by two different assays suggests seasonal cross-reactivity or interference.

The research uses different assays, chemiluminescent-immunoassay (CI) and immunofluorescent (IF), to compare plasma adrenocorticotropic hormone (ACTH) concentrations in ponies and suggests seasonal variations in results. This could possibly be due to seasonal cross-reactivity or interference.

Objective and Methods

• The main aim of the study was to compare the results of plasma adrenocorticotropic hormone (ACTH) analysis employing both CI and IF methods on samples collected in different seasons (autumn and spring), and to examine any possible cross-reactivity.
• The study used plasma from nonlaminitic ponies for analysis. The samples were analyzed concurrently using the IF and CI methods in autumn and the following spring.
• The IF method was calibrated using ROC curves and Youden indices to align it with the thresholds of the CI method. They examined the specificity of the assays using ACTH fragments and degradation products that were commercially available.
• The variations in the results of the assays implied possible seasonally dependent cross-reactivity or interference.

Results

• The study observed that the CI and IF methods yielded different results (P<0.001), with the mean differences indicating that the association between CI and IF results differed in autumn and spring. This supports the hypothesis of seasonally dependent cross-reactivity or interference.
• Good agreement was observed for binary interpretation in spring between IF and CI at certain thresholds, and in autumn between IF and CI at other thresholds.
• Of the ponies which provided both autumn and spring samples, 64% exceeded a particular CI threshold in autumn, but 70% of these were below the equivalent threshold the following spring without treatment.
• The CI assay showed apparent increases in ACTH following the addition of a certain peptide (CLIP – ACTH 18-39). They also found that degradation of ACTH during storage affected the assays differently.

Conclusions

• The study notes that a limited number of PPID cases were included, and the immunoreactivity of commercially available peptides may be different from their endogenous counterparts.
• The methods yielded different absolute values but the agreement for binary classification was good.
• The study suggests an altered pituitary secretome in autumn as likely responsible for affecting apparent ACTH values.