Use of a Granulocyte Immunofluorescence Assay Designed for Humans for Detection of Antineutrophil Cytoplasmic Antibodies in Dogs with Chronic Enteropathies

Abstract

BackgroundPerinuclear antineutrophil cytoplasmic antibodies (pANCA) previously have been shown to be serum markers in dogs with chronic enteropathies, with dogs that have food‐responsive disease (FRD) having higher frequencies of seropositivity than dogs with steroid‐responsive disease (SRD). The indirect immunofluorescence (IIF) assay used in previous publications is time‐consuming to perform, with low interobserver agreement.Hypothesis/ObjectivesWe hypothesized that a commercially available granulocyte IIF assay designed for humans could be used to detect perinuclear antineutrophil cytoplasmic antibodies in dogs.AnimalsForty‐four dogs with FRD, 20 dogs with SRD, 20 control dogs, and 38 soft‐coated wheaten terrier (SCWT) or SCWT‐cross dogs.MethodsA granulocyte assay designed for humans was used to detect pANCA, cANCA, and antinuclear antibodies (ANA), as well as antibodies against proteinase‐3 protein (PR‐3) and myeloperoxidase protein (MPO) in archived serum samples.ResultsSensitivity of the granulocyte assay to predict FRD in dogs was 0.61 (95% confidence interval (CI), 0.45, 0.75), and specificity was 1.00 (95% CI, 0.91, 1.00). A significant association was identified between positive pANCA or cANCA result and diagnosis of FRD (P < 0.0001). Agreement between the two assays to detect ANCA in the same serum samples from SCWT with protein‐losing enteropathy/protein‐losing nephropathy (PLE/PLN) was substantial (kappa, 0.77; 95% CI, 0.53, 1.00). Eight ANCA‐positive cases were positive for MPO or PR‐3 antibodies.Conclusions and Clinical ImportanceThe granulocyte immunofluorescence assay used in our pilot study was easy and quick to perform. Agreement with the previously published method was good.

Circulating Endocannabinoids as Diagnostic Markers of Canine Chronic Enteropathies: A Pilot Study

“…Accordingly, CRP was herein found high in the sera of only four out of 17 dogs, regardless of the final diagnosis, without differences among the dogs affected by different forms of CE, including those dogs affected by PLE. In one study (48), the seropositivity for pANCA discriminated IRE/NRE from other causes of diarrhea, with a sensitivity of 51% and a specificity of 83%, while in other studies the pANCA was found higher in sera of dogs with FRE than in sera of those affected by IRE/NRE (Se 61-62%; Sp 77-100%) (49,50). However, similar to serum CRP, seropositivity for pANCA can be also detected with other inflammatory conditions (51).…”

Clinical utility of currently available biomarkers in inflammatory enteropathies of dogs

“…Seropositivity for pANCA (and also pANCA titers) is higher in dogs with FRE (61%‐62%) than in those with IRE or NRE (0%‐37%) 95, 99, 100, 101. However, pANCA seropositivity is not specific for CIE and also can occur with other immune‐mediated, infectious, or neoplastic diseases 101, 102.…”

“…Perinuclear anti-neutrophilic cytoplasmic antibodies (pANCA) are serum autoantibodies against neutrophil granule components (eg, nuclear histone, proteinase 3, myeloperoxidase) and suspected crossreactivity with a gastrointestinal bacterial antigen 98 that can be detected by indirect immunofluorescence assays. 98,99 Seropositivity for pANCA (and also pANCA titers) is higher in dogs with FRE (61%-62%) than in those with IRE or NRE (0%-37%…”

Current diagnostics for chronic enteropathies in dogs

“…It is, therefore, prudent to always correlate the positivity of these tests with clinical findings. Of note, pANCA testing has recently been shown to correlate well with an indirect immunofluorescent method using human granulocytes as antigens, which is a clear advantage for further commercialization efforts 39 …”