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Frontiers in veterinary science2026; 13; 1884314; doi: 10.3389/fvets.2026.1884314

Reference interval for serum amyloid a in apparently healthy donkeys measured with a donkey-specific ELISA.

Abstract: Serum amyloid A (SAA) is a major acute phase protein used as an inflammatory biomarker in equine medicine. However, no universally accepted assay-specific reference interval (RI) exists for donkeys, and available literature remains limited, with only few studies using different donkey breeds and immunoassay platforms. This study aimed to establish an assay- and population-specific RI for serum SAA in apparently healthy donkeys using a species-specific double sandwich ELISA. Unassigned: Serum SAA concentrations were measured in 176 apparently healthy donkeys. RI estimation followed ASVCP/CLSI recommendations. The primary RI was calculated nonparametrically as the central 95% interval from all reference individuals, and two-sided 90% confidence intervals (CIs) for the reference limits were estimated nonparametrically. Data were inspected using Tukey's interquartile fences; however, high-end observations were not excluded from the primary RI analysis. Exploratory analyses were performed to assess the need for partitioning by sex, age, or breed. Unassigned: The nonparametric RI for serum SAA was 2.91-42.85 ng/mL. The 90% CI for the lower reference limit was 0.00-4.07 ng/mL, and the 90% CI for the upper reference limit was 35.49-67.10 ng/mL. Exploratory analyses did not support partitioning by sex, age, or breed. The relatively broad CI for the upper reference limit indicated imprecision, likely reflecting the influence of a minority of high-end observations. Unassigned: The low SAA concentrations observed in this study are consistent with findings reported in horses. Differences between this and previous donkey studies may reflect variation in assay platform, sample size, management conditions, physiological state, or population characteristics. Because SAA is a rapidly responsive acute phase protein, the proposed interval should be interpreted as an assay- and population-specific RI rather than as a disease-classification decision limit. Clinical interpretation should therefore always consider the individual donkey's clinical context.
Copyright © 2026 Perzyna, Bardet, Baranowska, Kiełbik, Puchalska, Pawliński and Witkowska-Piłaszewicz.
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Publication Date: 2026-07-13 PubMed ID: 42516301PubMed Central: PMC13402198DOI: 10.3389/fvets.2026.1884314Google Scholar: Lookup
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Summary

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Overview

  • This study established a reference interval (RI) for serum amyloid A (SAA) levels in healthy donkeys using a donkey-specific ELISA assay to improve the accuracy of inflammatory biomarker interpretation in equine medicine.
  • The research provides an assay- and population-specific baseline of SAA concentrations, allowing better clinical decision-making regarding inflammation in donkeys.

Background

  • Serum amyloid A (SAA): A major acute phase protein used as a biomarker for inflammation in horses and donkeys.
  • Importance of Reference Interval (RI): Clinicians rely on RIs to differentiate between healthy and diseased states based on biomarker levels.
  • Current Gap: No universally accepted, assay-specific RI for SAA in donkeys exists; previous studies used various breeds and assays, leading to inconsistent data.

Study Objectives and Design

  • To establish an assay- and population-specific RI for serum SAA in apparently healthy donkeys.
  • Used a species-specific double sandwich ELISA tailored for donkeys to measure serum SAA concentrations, ensuring assay specificity.
  • Sample included 176 apparently healthy donkeys, encompassing diverse ages, sexes, and breeds.
  • RI estimation followed established veterinary guidelines (ASVCP/CLSI), using nonparametric methods to determine the central 95% interval (2.5th to 97.5th percentile).
  • Two-sided 90% confidence intervals (CIs) were computed for the reference limits to assess statistical precision.
  • Data were inspected for outliers using Tukey’s interquartile fences, but high-end values were retained to reflect true population variance.
  • Exploratory analyses investigated whether RI partitioning was warranted based on sex, age, or breed differences.

Key Findings

  • The established nonparametric RI for serum SAA concentrations in donkeys was 2.91 to 42.85 ng/mL.
  • The 90% CI for the lower reference limit ranged from 0.00 to 4.07 ng/mL.
  • The 90% CI for the upper reference limit was wider, at 35.49 to 67.10 ng/mL, indicating some imprecision, likely due to a few high SAA values in the dataset.
  • No statistically significant need was found to partition the RI by sex, age, or breed, suggesting the interval applies broadly across these variables in donkeys.
  • The relatively low SAA concentrations align closely with previous findings observed in horses, reinforcing species similarities.

Interpretation and Significance

  • Differences between this study’s RI and those from earlier donkey research likely result from variations in assay platforms, sample sizes, donkey management, physiological conditions, or population profiles.
  • SAA is a highly dynamic acute phase protein that can rapidly increase during inflammation, so the presented RI should be viewed as an assay- and population-specific baseline rather than strict disease thresholds.
  • Clinical decisions should always consider the individual donkey’s health status and contextual factors in addition to serum SAA levels.
  • This donkey-specific assay and RI provide veterinarians with a more tailored and reliable tool to interpret SAA measurements for health monitoring and disease detection in donkeys.

Conclusion

  • The study successfully established a donkey-specific reference interval for serum amyloid A using a species-specific ELISA, improving the clinical utility of SAA as an inflammatory biomarker in donkeys.
  • The results emphasize the importance of assay- and population-specific RIs rather than relying on universal values derived from different species or assays.
  • The provided RI supports better differentiation of normal versus elevated SAA levels in healthy donkeys, enhancing disease monitoring and veterinary care.

Cite This Article

APA
Perzyna M, Bardet E, Baranowska K, Kiełbik P, Puchalska M, Pawliński B, Witkowska-Piłaszewicz O. (2026). Reference interval for serum amyloid a in apparently healthy donkeys measured with a donkey-specific ELISA. Front Vet Sci, 13, 1884314. https://doi.org/10.3389/fvets.2026.1884314

Publication

Frontiers in veterinary science
ISSN: 2297-1769
NlmUniqueID: 101666658
Country: Switzerland
Language: English
Volume: 13
Pages: 1884314
PII: 1884314

Researcher Affiliations

Perzyna, Maciej
  • Department of Large Animals Diseases and Clinic, Institute of Veterinary Medicine, Warsaw University of Life Sciences, Warsaw, Poland.
Bardet, Edith
  • Scientific Society of Veterinary Medicine Students, Warsaw University of Life Sciences, Warsaw, Poland.
Baranowska, Katarzyna
  • Scientific Society of Veterinary Medicine Students, Warsaw University of Life Sciences, Warsaw, Poland.
Kiełbik, Paula
  • Department of Large Animals Diseases and Clinic, Institute of Veterinary Medicine, Warsaw University of Life Sciences, Warsaw, Poland.
Puchalska, Maria
  • Department of Pathology and Veterinary Diagnostic, Institute of Veterinary Medicine, Warsaw University of Life Sciences, Warsaw, Poland.
Pawliński, Bartosz
  • Department of Large Animals Diseases and Clinic, Institute of Veterinary Medicine, Warsaw University of Life Sciences, Warsaw, Poland.
Witkowska-Piłaszewicz, Olga
  • Department of Large Animals Diseases and Clinic, Institute of Veterinary Medicine, Warsaw University of Life Sciences, Warsaw, Poland.

Conflict of Interest Statement

The author(s) declared that this work was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

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