Clinical Information

Antiphospholipid syndrome (APS) is a systemic autoimmune disease characterized by thrombosis and/or specific pregnancy-related death. Based on the 2006 revised Sapporo consensus classification criteria, the laboratory requirements for diagnosing APS include the presence of at least one of the following: lupus anticoagulant (LAC), anticardiolipin (aCL), and anti-beta-2 glycoprotein 1 (B2GP1) IgG or IgM antibodies.(1). To avoid overdiagnosis, and to exclude patients with transient antiphospholipid (aPL) levels, the APS guidance also recommends confirmation of any positive result at least 12 weeks after the initial evaluation. Of note, aPL antibodies also occur in patients with autoimmune diseases with significant prevalence in systemic lupus erythematosus (SLE) as well as other clinical manifestations (eg, heart valve disease, livedo reticularis, thrombocytopenia, nephropathy, and neurological) often associated with APS.(1-3) Thus, in addition to the 2006 APS guidance, the 2012 derivation and validation of the Systemic Lupus International Collaborating Clinics (SLICC) classification criteria for SLE recommends testing for the criteria aPL antibody tests as well as aCL IgA and anti-B2GP1 IgA.(2)

B2GP1 is a 326-amino acid protein synthesized by hepatocytes, endothelial cells, and trophoblast cells.(4) It contains 5 repetitive structures or "sushi domains," termed domain 1 through 5, for a combined molecular weight of 54?kDa for the protein.(4-6). Autoantibodies to B2GP1 may be detected by solid-phase immunoassays (SPA) and functional coagulation assays. Unlike LAC, the SPA provides quantitative measurements and antibody isotype class determinations that are important for risk assessment. Immunoassays for B2GP1 antibodies can be performed using either a composite substrate comprised of B2GP1 plus anionic phospholipid (ie, cardiolipin-dependent B2GP1) or B2GP1 alone. Antibodies detected using B2GP1 substrate without another phospholipid (direct assays) are referred to simply as "B2GP1 antibodies." Some B2GP1 antibodies are capable of inhibiting clot formation in functional coagulation assays that contain low concentrations of phospholipid cofactors.(6) Antibodies detected by functional coagulation assays are commonly referred to as LAC. Anti-B2GP1 antibodies associated with thromboembolic events target domain 1 of the molecule and are responsible for LAC (functional, phospholipid-dependent prolongation of the clotting time) and aCL-dependent B2GP1 antibody positivity.(7)

For detection of anti-B2GP1 IgG and IgM antibodies, the APS guidance advocates for the use of values above the 99th percentile of the laboratory’s population in the establishment of reference intervals for tests. While this recommendation may be used for anti-B2GP1 IgA immunoassays, there is no consensus for their determination.(6) For aCL IgG and IgM testing, the APS classification guidance recommends antibody cutoff values greater than 40 IgG phospholipid (GPL) or IgM PL (units traceable to the Harris standards for aCL antibody assays) or more than the 99th percentile for the testing laboratory’s population for positivity.(1) The use of cutoff values greater than 40 GPL or MPL units to define positivity is not applicable to all aCL antibody immunoassays as the threshold used to distinguish moderate-to-high positive from low positive results are test dependent.(4,5,7,8) In addition, the cutoff value used at the 99th percentile of a laboratory’s testing population may not be consistent with kits from the same manufacturer or 40 GPL units in the case of aCL antibodies.(5-7).

Thrombosis and obstetric complications are common clinical events in the general population and are not unique to APS; therefore, the presence of aPL antibodies is an absolute requirement for the diagnosis of definite APS.(1,5,7) Furthermore, aPL antibodies are heterogeneous with overlapping tendencies; the lack of aPL test harmonization or standardization requires the use of all 3 tests for optimal APS diagnosis.(1,5-7) The APL antibodies were traditionally determined using the classic enzyme-linked immunosorbent assay, with more diverse methods recently developed and adapted for clinical testing. Recognizing the analytical and diagnostic challenges associated with aPL antibody testing, initiatives to support assay harmonization and utilization, including the development of calibrators, test development, and validation efforts, as well as preanalytical, analytical, and postanalytical measures, have been published.(5-7) Based on these and other published studies, the interpretation and relevance of APL antibody tests are dependent on factors such as the type of aPL (LAC, aCL or anti-B2GP1), the source of cardiolipin and/or B2GP1, aPL antibody class (IgG, IgM or IgA) and level, as well as whether antibody positivity is single, double or triple.(1,5-8)

In conclusion, although the APS classification criteria were not established for routine clinical use, in the absence of formal diagnostic guidelines, these have widely been adopted to diagnose or assess risk for APS and the need for treatment or prophylaxis. Therefore, in clinical practice, if suspicion for disease is high but criteria aPL antibody tests are inconclusive or negative, deviation from the APS diagnostic criteria may be justified. This may include testing for non-criteria aPL antibody tests such as the aCL IgA and anti-B2GP1 IgA recommended in 2012 SLICC guidance for SLE and/or evaluation of anti-phosphatidylserine/prothrombin IgG and IgM autoantibodies, amongst others.(2,6,8-10)