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Veterinary clinical pathology2026; doi: 10.1111/vcp.70118

Evaluation of an Immunoturbidometric Assay for Measurement of Total Adiponectin Concentration in Equine Serum Samples.

Abstract: Total adiponectin (TA) is a marker of metabolic health in equids, with values below a critical decision point of < 7.9 μg/mL conferring increased risk of laminitis. Objective: To evaluate an IT assay to measure serum [TA] in equids and to assess its correlation with [high molecular weight adiponectin] (HMWA) measured by ELISA. Methods: Total serum adiponectin concentrations from 207 adult horses were measured using both an IT assay for TA and HMWA using a previously validated ELISA assay. Data were analyzed using Spearman correlation and Passing-Bablok regression. Intra- and inter-assay precision, linearity, lower limit of quantification (LLoQ), and freeze-thaw stability were assessed for the [TA] assay. Results: Mean (range) intra-assay coefficient of variation (CV%) was 2.3% (1.8%-2.8%); inter-assay, across-day CV% was 3.3% (3.25%-3.4%), and across reagent lot CV was 11.7% (9.0%-17.9%). Mean recovery upon dilution was 133.7% (122.6%-140.6%), and LLoQ 1.3 μg/mL. The median CV for 5 freeze-thaw cycles was 3.0% (range 1.0%-3.0%) with a median change in concentration of -11% (range -[9.9-17.1]%). A strong correlation between [TA] and [HMWA] is observed (ρ = 0.90 [95% confidence interval 0.86-0.92], p  [TA]. Conclusions: The turbidimetric immunoassay is sufficiently precise and reliable for measurement of [TA] in equids, especially around the critical decision point, and LLoQ was below the lowest clinical cut-off value. [TA] overall correlated well with [HMWA] in this population, although instances of higher [HMWA] than [TA] were noted at high [HMWA].
© 2026 The Author(s). Veterinary Clinical Pathology published by Wiley Periodicals LLC on behalf of American Society for Veterinary Clinical Pathology.
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Publication Date: 2026-07-22 PubMed ID: 42487462DOI: 10.1111/vcp.70118Google Scholar: Lookup
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Summary

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Overview

  • This study evaluates a new immunoturbidometric (IT) assay for measuring total adiponectin (TA) levels in horse serum, which is important for assessing metabolic health and the risk of laminitis.
  • The researchers compare the IT assay’s results with those from an established ELISA test for high molecular weight adiponectin (HMWA), aiming to validate the new method’s accuracy and reliability.

Background and Purpose

  • Adiponectin and Equine Health: Total adiponectin (TA) is an important biomarker indicating metabolic health in horses.
  • Clinical Relevance: TA levels below 7.9 μg/mL are linked to an increased risk of laminitis, a painful hoof condition.
  • Assay Evaluation: The study’s main goal was to test the performance of an immunoturbidometric (IT) assay to measure total adiponectin in horse serum samples.
  • Comparison Metric: The researchers also wanted to assess how TA measured by this assay correlates with high molecular weight adiponectin (HMWA) measured via ELISA, considering HMWA is another relevant adiponectin form related to metabolic health.

Methods

  • Sample Population: Serum samples were collected from 207 adult horses to ensure a broad representative dataset.
  • Testing Procedures:
    • TA concentrations were measured using the new immunoturbidometric assay.
    • HMWA concentrations were measured using a previously validated enzyme-linked immunosorbent assay (ELISA).
  • Data Analysis:
    • Spearman correlation was used to assess the relationship between TA and HMWA values.
    • Passing-Bablok regression analysis was conducted to evaluate equivalency and bias between the two assays.
  • Assay Performance Parameters Evaluated:
    • Intra-assay precision (repeatability within the same test run)
    • Inter-assay precision (repeatability across different runs/days and reagent lots)
    • Linearity (accuracy across a range of dilutions)
    • Lower Limit of Quantification (LLoQ) — lowest reliable detection checkpoint
    • Freeze-thaw stability to evaluate sample integrity after multiple freeze-thaw cycles

Results

  • Precision:
    • Intra-assay coefficient of variation (CV) averaged 2.3%, indicating high repeatability within runs.
    • Inter-assay CV was 3.3% across days, maintaining low variability.
    • Variability across different reagent lots was higher (mean CV ~11.7%), highlighting some batch differences.
  • Linearity and Recovery:
    • The assay showed a recovery of about 134% upon dilution, which is somewhat higher than ideal and suggests an overestimation in diluted samples.
  • Lower Limit of Quantification (LLoQ):
    • The assay could reliably detect adiponectin down to 1.3 μg/mL, well below the critical clinical threshold of 7.9 μg/mL important for risk assessment.
  • Freeze-Thaw Stability:
    • Median coefficient of variation across 5 freeze-thaw cycles was only 3.0%, indicating stable results.
    • There was a median concentration reduction of about 11%, indicating some loss but likely within acceptable limits for clinical use.
  • Correlation Between TA (IT assay) and HMWA (ELISA):
    • A strong positive correlation was observed (Spearman ρ = 0.90), confirming the assays generally provide consistent results.
    • In 18% of samples, HMWA levels were higher than total adiponectin, an unexpected finding suggesting some differences in what each assay measures, especially with high HMWA concentrations.

Conclusions and Implications

  • The new immunoturbidometric assay is precise and reliable for measuring total adiponectin in horse serum.
  • Its sensitivity and detection limits make it especially suitable for clinical use, including identifying animals at risk for laminitis due to low adiponectin levels.
  • The strong correlation with an established ELISA assay for HMWA supports its validity, despite some discrepancies at high concentrations.
  • This assay offers a practical alternative that may be easier or faster to implement than ELISA in routine veterinary diagnostics.
  • Further investigation might be needed to understand the instances where HMWA exceeds total adiponectin measurements and how these relate to clinical outcomes.

Cite This Article

APA
Nolen-Walston RD, Lowndes CR, Kulp JC, Luethy D, Stefanovski D, Van Eps AW. (2026). Evaluation of an Immunoturbidometric Assay for Measurement of Total Adiponectin Concentration in Equine Serum Samples. Vet Clin Pathol. https://doi.org/10.1111/vcp.70118

Publication

Veterinary clinical pathology
ISSN: 1939-165X
NlmUniqueID: 9880575
Country: United States
Language: English

Researcher Affiliations

Nolen-Walston, Rose D
  • Department of Clinical Studies-New Bolton Center, School of Veterinary Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, USA.
Lowndes, Caitrin R
  • Department of Clinical Studies-New Bolton Center, School of Veterinary Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, USA.
Kulp, Jeaneen C
  • Department of Clinical Studies-New Bolton Center, School of Veterinary Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, USA.
Luethy, Daniela
  • Department of Clinical Studies-New Bolton Center, School of Veterinary Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, USA.
Stefanovski, Darko
  • Department of Clinical Studies-New Bolton Center, School of Veterinary Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, USA.
Van Eps, Andrew W
  • Department of Clinical Studies-New Bolton Center, School of Veterinary Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, USA.

Grant Funding

  • the Laminitis Research Fund, University of Pennsylvania

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