Abstract
The chemiluminescence (CL) and immunofluorescence (IF) assays yield different results for basal adrenocorticotropin hormone concentrations [ACTH] in pony plasma. It is unclear whether this difference also occurs in basal samples from horses or samples from ponies following thyrotropin-releasing hormone (TRH) stimulation. To compare the results of [ACTH] analysis by CL and IF methods in basal samples from horses and pony samples following TRH stimulation. Method comparison. Plasma [ACTH] was measured concurrently using CL and IF methods in 12 ponies (basal and post-TRH stimulation) in November and basal samples from horses (n=45; November and May). CL and IF methods yielded different results (P<.01). The median difference (CL-IF) (95% CI) for ponies was 5.9 (0.1-7.5)pg/mL at baseline and 227.9 (61-1001)pg/mL post TRH; and horses 1.9 (1.1-5.4)pg/mL in November and 9.4 (8.2-11.5)pg/mL in May, at baseline. Correlation was good in ponies at baseline (R=0.80, P=.003) but not post-TRH, and good in horses in November and May (R=0.68 and 0.71, P<.001). Bland-Altman analysis demonstrated moderate bias and wide 95% limits of agreement (95% LOA) in ponies at baseline (bias 5.5pg/mL; 95% LOA -9.9 to 20.9pg/mL) and horses in May (bias 10.6pg/mL; 95% LOA -9 to 30.3pg/mL) and very large bias and wide 95% LOA in ponies post-TRH (bias 477pg/mL; 95% LOA -633 to 1587pg/mL). Using CL cut-offs of >29 and >110pg/mL, agreement was moderate (ƙ=0.67) and very good (ƙ=0.82) for binary classification of PPID in ponies at baseline and post-TRH; and good (ƙ=0.73) for horses in November, but poor (ƙ=0.40) in May. Limited numbers of horses with [ACTH] above threshold values. The assays yielded different absolute values, particularly in post-TRH samples from ponies, suggesting TRH stimulates secretion of cross-reacting peptides other than ACTH. Agreement for binary classification for PPID was moderate to good, except in basal samples from horses in May.
Highlights
Analysis of plasma adrenocorticotropin hormone concentration [ACTH], both at baseline and following thyrotropinreleasing hormone (TRH) stimulation, is used to aid the diagnosis of equine pituitary pars intermedia dysfunction (PPID) [1]
Correlation was good in ponies at baseline (R=0.80, p=0.003) but not post TRH, and good in horses in November and May (R=0.68 and 0.71, p
The post TRH stimulation test [ACTH] cut-off values are an ongoing area of research, with current cut-offs indicating a wide range of equivocal values [6]
Summary
Analysis of plasma adrenocorticotropin hormone concentration [ACTH], both at baseline and following thyrotropinreleasing hormone (TRH) stimulation, is used to aid the diagnosis of equine pituitary pars intermedia dysfunction (PPID) [1]. The TRH stimulation test can be used to assist diagnosis of PPID where baseline [ACTH] is equivocal or negative and PPID is suspected. Plasma [ACTH] can vary in individuals and TRH stimulation tests are thought to reduce the relative impact of confounding factors by maximally and stimulating the pars intermedia of the pituitary gland [2]. Adjusted CL reference ranges for baseline [ACTH] are widely used, and more recently seasonally adjusted post TRH stimulation [ACTH] cut-offs have been reported [4,5]. The post TRH stimulation test [ACTH] cut-off values are an ongoing area of research, with current cut-offs indicating a wide range of equivocal values [6]
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