Background/Purpose:

Identification of serum anti-neutrophil cytoplasmic antibodies (ANCA) for the detection of ANCA-associated vasculitis (AAV) is often performed by screening with indirect immunofluorescence (IF), followed by testing of IF-positive samples for anti-proteinase-3 (PR3) and anti-myeloperoxidase (MPO) antibodies.  Many institutions, including ours, analyze all samples by IF and anti-PR3/anti-MPO assays simultaneously.  This strategy can identify patients with detectable anti-PR3 or anti-MPO antibodies despite a negative IF (IF-/Ab+); however, the significance of this discordant result is not clear.  We attempted to determine whether IF-/Ab+ results identified any cases of clinically meaningful systemic vasculitis.

Methods:

We conducted a retrospective chart review of all patients tested for serum ANCAs at a large academic center between January 2011 and May 2013 who had a negative IF but detectable anti-PR3 or anti-MPO antibodies (IF-/Ab+).  IF was performed on serum diluted 1:20, with cytoplasmic or perinuclear patterns considered positive.  Atypical patterns were excluded.  Anti-PR3 and anti-MPO assays were performed by immunoassay.  AAV diagnosis was based on the European Medicines Agency vasculitis algorithm. 

Results:

Of 2345 samples tested for ANCAs, 49 samples (2.1%), derived from 38 patients, contained a detectable anti-PR3 or anti-MPO antibody despite a negative IF.  By comparison, 1950 samples (83%) had both negative IF and antibody assays, and 123 samples (5.2%), derived from 68 patients, had both a positive IF and a detectable anti-PR3 (44), anti-MPO antibody (77), or both antibodies (2).  Of 76 samples with detectable anti-PR3 antibodies, 29 (38%) had a negative IF, while 20 of 115 (17%) anti-MPO-positive samples had a negative IF.

We identified only one case in which a patient with an IF-/Ab+ result was subsequently diagnosed with AAV.  A repeat test 1 month later returned IF+/Ab+.  Eleven of the IF-/Ab+ cases (29%) represented previously diagnosed and treated AAV, all with positive IF and antibody tests prior to treatment.  The majority of patients with IF-/Ab+ results (20/38 (52%)) had non-vasculitic immunologic disorders, including SLE (5) inflammatory bowel disease (3), autoimmune hepatitis (1), Hashimoto’s thyroiditis (1), relapsing polychondritis (1), pyoderma gangrenosum (1), hepatitis C (1), and graft versus host disease (1).  In the comparator group, 42 of 68 (62%) patients with IF+/Ab+ results were diagnosed with AAV (anti-PR3 17, anti-MPO 25).

Conclusion:

A positive anti-PR3 or anti-MPO assay in the absence of positive ANCA IF may rarely lead to a new diagnosis of systemic vasculitis; however, this immune profile can be observed in patients with previously known seropositive (IF+/Ab+) AAV.  If one believes that serial ANCA testing is of value in monitoring clinical activity and response to therapy, our data suggest that both IF and specific antibody tests should be monitored, though the utility of this practice is controversial.  Further investigation into the clinical implications of persistent antibody-positivity with negative immunofluorescence is warranted.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/the-significance-of-anti-myeloperoxidase-and-anti-proteinase-3-antibodies-in-the-absence-of-anti-neutrophil-cytoplasmic-antibody-immunofluorescence-positivity/