The immune system produces antibodies to neutralize foreign invaders. In autoimmune conditions, the body mistakenly creates autoantibodies that target its own healthy cells. Antineutrophil Cytoplasmic Antibodies (ANCA) are a specific type of autoantibody that targets components within neutrophils, the most common infection-fighting white blood cell, and ANCA testing aids in diagnosing various systemic autoimmune diseases.
Understanding Classic ANCA Patterns
The initial method for detecting ANCA involves an Indirect Immunofluorescence Assay (IFA), where a patient’s serum is applied to a slide containing ethanol-fixed neutrophils. This technique reveals the location where the autoantibodies bind, resulting in characteristic glowing patterns visible under a fluorescent microscope. The two primary, or classic, patterns identified by this method are the Cytoplasmic ANCA (C-ANCA) and the Perinuclear ANCA (P-ANCA).
The C-ANCA pattern appears as a granular, diffuse glow spread throughout the neutrophil’s cytoplasm, strongly associated with antibodies targeting the enzyme Proteinase 3 (PR3). The classic P-ANCA pattern shows a dense, ring-like glow concentrated around the nucleus of the cell, overwhelmingly linked to antibodies directed against the enzyme Myeloperoxidase (MPO). The presence of these classic ANCA patterns is highly relevant to diagnosing small-vessel vasculitis conditions, such as Granulomatosis with Polyangiitis (PR3-ANCA) and Microscopic Polyangiitis (MPO-ANCA).
Defining Atypical P-ANCA
The term “Atypical P-ANCA” (or A-ANCA) describes a result showing the P-ANCA staining pattern on the immunofluorescence test, but which is not caused by the typical target, Myeloperoxidase (MPO). Although the visual result appears perinuclear, the autoantibodies target other, less common proteins within the neutrophil. This means the IFA is positive for the P-ANCA pattern, but subsequent specific testing for MPO will be negative.
The perinuclear appearance of these atypical antibodies is often an artifact of the ethanol fixation process used in the IFA test. When the neutrophil is fixed with ethanol, many cytoplasmic antigens, which are positively charged, relocate and cluster around the negatively charged nuclear membrane. This relocation causes the antibody binding to appear as a perinuclear ring, even though the target protein may have originally been spread throughout the cytoplasm.
The non-classic antigens targeted by Atypical P-ANCA are diverse and include proteins such as lactoferrin, cathepsin G, elastase, and bactericidal/permeability-increasing protein (BPI). These antibodies are distinct from classic MPO and PR3 antibodies and are not typically associated with severe systemic vasculitis diseases. The identification of an Atypical P-ANCA pattern directs clinicians to consider a different set of autoimmune conditions.
Clinical Significance in Autoimmune Disease
The clinical relevance of Atypical P-ANCA lies primarily outside the classic ANCA-associated vasculitides. It serves as a serological marker for chronic inflammatory conditions, most notably Inflammatory Bowel Disease (IBD). Atypical P-ANCA is particularly prevalent in patients with Ulcerative Colitis (UC), with studies showing a positive result in 40% to 80% of patients.
In the context of IBD, Atypical P-ANCA helps distinguish UC from Crohn’s Disease (CD), the other major form of IBD, where the antibody is less common. Its presence can support a clinical diagnosis of UC, especially when the disease presentation is ambiguous or overlaps with CD. The presence of this autoantibody has also been associated with the extent of colitis, sometimes correlating with more widespread involvement of the colon.
Beyond IBD, Atypical P-ANCA is also observed in other autoimmune disorders that affect the liver and biliary system, such as Primary Sclerosing Cholangitis (PSC) and Autoimmune Hepatitis (AIH). Although it is not a standalone diagnostic test, the result offers valuable supporting evidence for the clinician alongside other laboratory or imaging results. The presence of these non-MPO/PR3 antibodies suggests a different immune pathway is activated compared to the one seen in classic vasculitis.
How Atypical P-ANCA is Tested
Identifying an Atypical P-ANCA result requires a two-step laboratory approach to ensure an accurate interpretation. The first step involves the screening test, the Indirect Immunofluorescence Assay (IFA), where the patient’s serum is incubated with fixed neutrophils. If autoantibodies are present, they will bind and create the glowing P-ANCA pattern visible under the microscope, indicating a positive result.
The second, confirming step uses antigen-specific Enzyme-Linked Immunosorbent Assay (ELISA) tests. This test is designed to check specifically for the presence of antibodies against the two main classic antigens: Myeloperoxidase (MPO) and Proteinase 3 (PR3). If the initial IFA screen is positive for the P-ANCA pattern, but the subsequent MPO and PR3 ELISA tests are both negative, the result is then confirmed to be Atypical P-ANCA.
The combination of a positive P-ANCA pattern on IFA and negative MPO/PR3 results on ELISA indicates that the autoantibodies are targeting one of the non-classic antigens, such as lactoferrin or elastase. This two-step process is necessary because the visual IFA pattern alone cannot differentiate between the clinically distinct classic MPO-ANCA (associated with vasculitis) and the Atypical P-ANCA (often associated with IBD). By ruling out MPO and PR3 as the target, the laboratory provides the specific information necessary for the clinician to interpret the result in the proper clinical context.