IgA anti-β2GPI Research Articles

IgA Anti-β2-Glycoprotein I Antibodies as Markers of Thrombosis and Severity in COVID-19 Patients.

Patients with COVID-19 may develop a hypercoagulable state due to tissue and endothelial injury, produced by an unbalanced immune response. Therefore, an increased number of thromboembolic events has been reported in these patients. The aim of this study is to investigate the presence of antiphospholipid antibodies (aPL) in COVID-19 patients, their role in the development of thrombosis and their relationship with the severity of the disease. In this retrospective study, serum samples from 159 COVID-19 patients and 80 healthy donors were analysed for the presence of aPL. A total of 29 patients (18.2%) and 14 healthy donors (17.5%) were positive for aPL. Nineteen COVID-19 patients (12%) but no healthy donor presented a positive percentage of the IgA isotype aPL. IgA anti-β2-glycoprotein I antibodies (anti-β2GPI) were the most frequent type (6.3%) in patients but was not detected in any healthy donor. The positivity of this antibody was found to be significantly elevated in patients with thromboembolic events (25% vs. 5%, p = 0.029); in fact, patients with positive IgA anti-β2GPI had an incidence of thrombosis over six times higher than those who had normal antibody concentrations [OR (CI 95%) of 6.67 (1.5-30.2), p = 0.014]. Additionally, patients with moderate-severe disease presented a higher aPL positivity than patients with mild disease according to the Brescia (p = 0.029) and CURB-65 (p = 0.011) severity scales. A multivariate analysis showed that positivity for IgA anti-β2GPI is significantly associated with disease severity measured by CURB-65 [OR (CI 95%) 17.8 (1.7-187), p = 0.0016]. In conclusion, COVID-19 patients have a significantly higher positive percentage of the IgA isotype aPL than healthy donors. IgA anti-β2GPI antibodies were the most frequently detected aPL in COVID-19 patients and were associated with thrombosis and severe COVID-19 and are thus proposed as a possible marker to identify high-risk patients.

Prevalence, Domain Specificity, and Residue Specificity of IgG, IgM, and IgA Anti-β2GPI Antibodies in Tetra-Positive Antiphospholipid Syndrome Patients: Identification of Two Non-Overlapping Epitopes in Domain I of β2GPI

Background. Risk stratification of antiphospholipid syndrome (APS) patients involves several laboratory tests, including lupus anticoagulant (LAC), anti-cardiolipin antibodies (anti-CL), anti-β2GPI antibodies (anti-β2GPI), and, more recently, anti-phosphatidylserine/prothrombin antibodies (anti-PS/PT). Patients with positive results for all four tests, known as tetra-positive APS patients, are considered at high risk for thrombotic recurrence. β2GPI is a protein composed of five domains: DI, DII, DIII, DIV, and DV, arranged in a flexible elongated structure. Previous studies have demonstrated that DI is frequently targeted by IgG in thrombotic APS patients. However, the prevalence, domain specificity, and residue specificity of the three different isotypes of anti-β2GPI antibodies in tetra-positive APS patients remain unknown. Objective. To establish the prevalence, domain specificity, and residue specificity of IgG, IgM, and IgA anti-β2GPI antibodies in tetra-positive APS patients. Methods. Plasma samples from 32 clinically confirmed tetra-positive APS patients and 8 healthy controls were analyzed for levels of IgG, IgM, and IgA using an enzyme-linked immunosorbent assay (ELISA) with immobilized β2GPI as the antigen. Domain and residue mapping were investigated using five deleted domain variants and eight single-point mutations. Mutants lacking specific disulfide bonds were used to assess whether the epitopes targeted by the antibodies are linear or conformational. Results. Among the 32 patients tested, 31 (96%) had IgG, 18 (56%) had IgM, and 18 (56%) had IgA anti-β2GPI antibodies. Ten patients (31%) had all three isotypes, 7 (22%) had IgG and IgM, and 8 (25%) had IgG and IgA. Six patients (19%) had only IgG, 1 had only IgM, and none had only IgA. The removal of DI from the β2GPI molecule led to an almost complete loss of binding (~80%) for the majority of IgG antibodies. There was a lesser but still significant reduction in binding when DII and DV were removed. However, no loss of binding was observed with variants lacking DIII and DIV. Notably, IgA antibodies behaved similarly to IgG but differently from IgM, as their reactivity towards DI and DV was significantly attenuated. To investigate the contribution of specific residues to binding, charged residues K19, E23, E26, E27, R39, R43, K44, and K59 in DI were individually replaced with alanine (A). The most important residue for IgG and IgA antibodies was R43, followed by R39 and K44. In contrast, most residues contributed to optimal binding for IgM antibodies, suggesting a less specific epitope definition. Interestingly, the mutations R39A and K44A often led to loss of binding in conjunction with the mutation R43A, but rarely simultaneously, suggesting the existence of two distinct epitopes for anti-DI antibodies-one located north and one south of R43. Finally, IgG, IgM, and IgA antibodies did not react against the mutant C32S/C60S but showed good reactivity against the mutant C288S/C326S, indicating that epitopes in DI are conformational and require the integrity of the disulfide bond 32-60. Conclusions. In tetra-positive APS patients, IgG anti-β2GPI antibodies are the most prevalent isotype, followed by IgA and IgM. The IgG and IgA isotypes exhibit a pronounced preference for DI, while the IgM isotype demonstrates lower specificity, which aligns with IgM's role as initial responders in infections. Notably, our research revealed that anti-DI antibodies are diverse, as they recognize epitopes within DI that are closely related but physically distinct. Ongoing studies aim to further characterize these antibody subtypes and understand their significance in APS. This knowledge may inform improved diagnostic and therapeutic approaches for APS patients.

Longitudinal Analysis of Anti-cardiolipin and Anti-β2-glycoprotein-I Antibodies in Recent-Onset Systemic Lupus Erythematosus: A Prospective Study in Swedish Patients.

Background: Anti-phospholipid syndrome (APS) and systemic lupus erythematous (SLE) are autoimmune disorders that often co-occur. Anti-phospholipid antibodies (aPL) are typical of both conditions and may be associated with vascular events and pregnancy-related morbidities. Whereas, aPL-screening is mandatory for individuals with suspected SLE, the clinical value of longitudinal aPL analyses in established SLE is unclear.Methods: We investigated the occurrence and variation of IgG/IgA/IgM anti-cardiolipin (aCL) and anti-β2-glycoprotein-I (anti-β2GPI) antibodies, using both the manufacturer's cut-off and a cut-off based on the 99th percentile of 400 apparently healthy donors, in recent-onset SLE. Furthermore, we evaluated the relationships between aPL levels and SLE/APS manifestations, as well as the pharmacotherapy. Patients with SLE who met validated classification criteria were included in this prospective study (N = 54). Samples were obtained at 0, 6, 12, 24, 36, 48, 60, 72, 84, and 96 months after SLE diagnosis.Results: Depending on the cut-off applied, 61.1 or 44.4% showed a positive result for at least one aPL isotype or the lupus anticoagulant test over time. Median values for all six aPL isotypes numerically decreased from inclusion to last follow-up, but none of the isotypes met statistical significance. Seroconversion (from positive to negative, or the opposite direction) was occasionally seen for both aCL and anti-β2GPI. IgA and IgM anti-β2GPI were the most common isotypes, followed by IgM aCL. Presence of IgG aCL associated significantly with myocardial infarction and miscarriage, and IgG/IgA anti-β2GPI with miscarriage.Conclusion: aPL were common during the first years of SLE. Even though the levels fluctuated over time, the patients tended to remain aPL positive or negative. Repeated aPL testing in the absence of new symptoms seems to be of uncertain value in patients with recent-onset SLE.

Elevated IgA antiphospholipid antibodies in healthy pregnant women in Sudan but not Sweden, without corresponding increase in IgA anti-β2 glycoprotein I domain 1 antibodies

ObjectiveThe role of antiphospholipid antibodies (aPL) during apparently normal pregnancy is still unclear. IgA aPL are prevalent in populations of African origin. Our aim was to measure all isotypes of anticardiolipin (anti-CL) and anti–β2 glycoprotein I (anti-β2GPI) in healthy pregnant and non-pregnant women of different ethnicities.MethodsHealthy Sudanese pregnant women (n = 165; 53 sampled shortly after delivery), 96 age-matched Sudanese female controls and 42 healthy pregnant and 249 non-pregnant Swedish women were included. IgA/G/M anti-CL and anti-β2GPI were tested at one time point only with two independent assays in Sudanese and serially in pregnant Swedes. IgA anti-β2GPI domain 1 and as controls IgA/G/M rheumatoid factor (RF), IgG anti–cyclic citrullinated peptide 2 (anti-CCP2) and anti–thyroid peroxidase (anti-TPO) were investigated in Sudanese females.ResultsPregnant Sudanese women had significantly higher median levels of IgA anti-CL, IgA anti-β2GPI (p < 0.0001 for both antibodies using two assays) and IgM anti-β2GPI (both assays; p < 0.0001 and 0.008) compared with non-pregnant Sudanese. IgA anti-CL and anti-β2GPI occurrence was increased among Sudanese pregnant women compared with national controls. No corresponding increase during pregnancy was found for IgA anti-β2GPI domain 1 antibodies. Both IgG anti-CL and IgG control autoantibodies decreased during and directly after pregnancy among Sudanese. Serially followed Swedish women showed no changes in IgA aPL, whereas IgG/M anti-CL decreased.ConclusionsIgA aPL are increased in Sudanese but not in Swedish women, without corresponding increase in IgA domain 1. Whether due to ethnicity and/or environmental influences the occurrence of IgA aPL during Sudanese pregnancies, and its clinical significance, is yet to be determined.

The Impact of Systemic Lupus Erythematosus on the Clinical Phenotype of Antiphospholipid Antibody–Positive Patients: Results From the AntiPhospholipid Syndrome Alliance for Clinical Trials and InternatiOnal Clinical Database and Repository

Although systemic lupus erythematosus (SLE) is the most common autoimmune disease associated with antiphospholipid antibodies (aPL), limited data exist regarding the impact of SLE on the clinical phenotype of aPL-positive patients. The primary objective of this study was to compare the clinical, laboratory, and treatment characteristics of aPL-positive patients with SLE with those of aPL-positive patients without SLE. A secure web-based data capture system was used to store patient demographic characteristics and aPL-related clinical and laboratory characteristics. Inclusion criteria included positive aPL according to the updated Sapporo classification criteria. Antiphospholipid antibody-positive patients fulfilling the American College of Rheumatology criteria for the classification of SLE ("aPL with SLE") and those with no other autoimmune diseases ("aPL only") were included in the analysis. Six hundred seventy-two aPL-positive patients were recruited from 24 international centers; 426 of these patients did not have other autoimmune disease, and 197 had SLE. The frequency of thrombocytopenia, hemolytic anemia, low complement levels, and IgA anti-β2 -glycoprotein I (anti-β2 GPI) antibodies was higher in the aPL-positive patients with SLE, whereas the frequency of cognitive dysfunction and IgG anti-β2 GPI antibodies was higher in the aPL-only group. The frequency of arterial and venous thromboses (including recurrent) as well as pregnancy morbidity was similar in the 2 groups. The prevalence of cardiovascular disease risk factors at the time of entry into the registry entry did not differ between the 2 groups, with the exception of current smoking, which was more frequent in aPL-positive patients with SLE. Although the frequencies of thrombosis and pregnancy morbidity are similar in aPL-positive patients with and those without SLE, the diagnosis of SLE in patients with persistently positive aPL is associated with an increased frequency of thrombocytopenia, hemolytic anemia, low complement levels, and positive IgA anti-β2 GPI antibodies.

Immunoglobulin A anti-phospholipid antibodies in Swedish cases of systemic lupus erythematosus: associations with disease phenotypes, vascular events and damage accrual.

Immunoglobulin (Ig) G- and IgM-class anti-cardiolipin antibodies (aCL) and lupus anti-coagulant (LA) are included in the 1997 update of the American College of Rheumatology (ACR-97) systemic lupus erythematosus (SLE) criteria. Despite limited evidence, IgA-aCL and IgA anti-β2 -glycoprotein-I (anti-β2 GPI) were included in the 2012 Systemic Lupus International Collaborating Clinics criteria. The present study aimed to evaluate IgG-/IgA-/IgM-aCL and anti-β2 GPI occurrence in relation to disease phenotype, smoking habits, pharmacotherapy, anti-phospholipid syndrome (APS) and organ damage among 526 Swedish SLE patients meeting ACR-97. Patients with rheumatoid arthritis (n=100), primary Sjögren's syndrome (n=50) and blood donors (n=507) served as controls. Anti-phospholipid antibodies (aPL) were analysed by fluoroenzyme-immunoassays detecting aCL/anti-β2 GPI. Seventy-six (14%) SLE cases fulfilled the Sydney APS-criteria, and ≥1 aCL/anti-β2 GPI isotype (IgG/IgA/IgM) occurred in 138 SLE patients (26%). Forty-five (9%) of the SLE cases had IgA-aCL, 20 of whom (4%) lacked IgG-/IgM-aCL. Seventy-four (14%) tested positive for IgA anti-β2 GPI, 34 (6%) being seronegative regarding IgG/IgM anti-β2 GPI. Six (1%) had APS manifestations but were seropositive regarding IgA-aCL and/or IgA anti-β2 GPI in the absence of IgG/IgM-aPL and LA. Positive LA and IgG-aPL tests were associated with most APS-related events and organ damage. Exclusive IgA anti-β2 GPI occurrence associated inversely with Caucasian ethnicity [odds ratio (OR)=0·21, 95% confidence interval (CI)=0·06-0·72) and photosensitivity (OR=0·19, 95% CI=0·05-0·72). Nephritis, smoking, LA-positivity and statin/corticosteroid-medication associated strongly with organ damage, whereas hydroxychloroquine-medication was protective. In conclusion, IgA-aPL is not rare in SLE (16%) and IgA-aPL analysis may have additional value among SLE cases with suspected APS testing negative for other isotypes of aPL and LA.

Live Birth Pregnancy Outcome after First In Vitro Fertilization Treatment in a Patient with Systemic Lupus Erythematosus and Isolated High Positive IgA Anti-β2glycoprotein I Antibodies: A Case Report.

IgA anti-β2glycoprotein I antibodies (IgA-anti-β2GPI) seems to be the most prevalent isotype in patients with Systemic Lupus Erythematosus (SLE) with a significant association to thrombotic events. Both SLE and antiphospholipid syndrome (APS) can be associated with implantation failure, fetal loss and obstetric complications. Recent reports highlight the clinical value of IgA-anti-β2GPI determination in supporting in vitro fertilization (IVF) treatment and IVF pregnancy outcomes. We report a 36-year-old female diagnosed with SLE, endometriosis and unexplained infertility. Conventional APS markers were consistently negative: anti-cardiolipin (aCL) and anti-β2GPI: IgG/IgM. She was then tested with reports of repeatedly high IgA-anti-β2GPI and tested positive from 2014 after IgA (aCL; anti-β2GPI) were established in our APS diagnostic panel. She underwent successful first IVF procedure with a 30 week live birth pregnancy outcome. During the follow up no lupus flare, thrombosis or ovarian hyperstimulation syndrome were registered. Serum IgA anti-β2GPI and anti-dsDNA levels declined statistically significant during the second and third trimester. Titres of IgA-anti-β2GPI remained lower postpartum as well. This case highlights the clinical importance of IgA-anti-β2GPI testing for family planning, assisted reproduction and pregnancy in women with SLE and/or APS.

Differential assay reactivity of immunglobulin A anti-ß2 glycoprotein I antibodies: implications for the clinical interpretation of antiphospholipid antibody testing.

The routine measurement of IgA anticardiolipin (aCL) and IgA anti-β2 glycoprotein I (anti-β2 GPI) antibodies remain controversial despite several studies demonstrating an association with thromboembolic disease in patients with systemic lupus erythematosus (SLE) and antiphospholipid syndrome (APS). This controversy may be a contributing factor for the current under use of IgA antiphospholipid antibodies. We aimed to investigate the nature of discrepant IgA anti-β2 GPI reactivity to help define the diagnostic value of IgA antiphospholipid antibodies. Four sera selected from SLE/APS patients and positive for antiphospholipid antibodies but having discrepant IgA anti-β2 GPI reactivity on two commercial assays were studied. IgA antibodies were affinity purified to investigate anti-β2 GPI reactivity. Column wash through and eluent fractions were tested on both IgA anti-β2 GPI assays. Results were normalized to total protein. Assay conjugates and standards from the discrepant assays were interchanged. The diseased samples were strongly positive in one assay [144-388 IgA antiphospholipid (APL) units] and negative or weakly positive in another assay (9.9-53 APL units). IgA eluents from IgA anti-β2 GPI positive samples reacted 10 times stronger on the reactive assay. When normalized to protein content, the eluents showed no cross-reactivity for IgG or IgM anti-β2 GPI antibodies, confirming IgA isotype specificity. Conjugate interchange confirmed that both assays bound IgA anti-β2 GPI antibodies, but the anti-IgA conjugate from the reactive assay was 4 times stronger, suggesting that its ability to detect IgA anti-β2 GPI antibodies was partially dependent on the anti-IgA conjugate and calibration. These results confirm not only the presence of IgA anti-β2 GPI antibodies in the selected patient samples but also highlight an IgA conjugate issue for the unreactive assay, causing an underestimation of IgA anti-β2 GPI. This finding may assist in the ongoing standardization efforts of APS antibody testing. In addition, conclusions from published clinical studies may need to be revised as some assays may understate IgA significance.

AB0558 What is the Prevalence of NON Criteria Antiphospholipid Antibodies in Patients with Antiphospholipid Syndrome?

BackgroundIncreasing interest has focussed upon assays which are not currently included in the Antiphospholipid syndrome (APS) classification criteria to detect antibodies directed against other phospholipids (PL), PL binding proteins and/or...

Value of Isolated IgA Anti–β2‐Glycoprotein I Positivity in the Diagnosis of the Antiphospholipid Syndrome

To examine the prevalence of isolated IgA anti-β2 -glycoprotein I (anti-β2 GPI) positivity and the association of these antibodies, and a subgroup that bind specifically to domain IV/V of β2 GPI, with clinical manifestations of the antiphospholipid syndrome (APS) in 3 patient groups and to evaluate the pathogenicity of IgA anti-β2 GPI in a mouse model of thrombosis. Patients with systemic lupus erythematosus (SLE) from a multiethnic, multicenter cohort (LUpus in MInorities, NAture versus nurture [LUMINA]) (n = 558), patients with SLE from the Hopkins Lupus Cohort (n = 215), and serum samples referred to the Antiphospholipid Standardization Laboratory (APLS) (n = 5,098) were evaluated. IgA anti-β2 GPI titers and binding to domain IV/V of β2 GPI were examined by enzyme-linked immunosorbent assay (ELISA). CD1 mice were inoculated with purified IgA anti-β2 GPI antibodies, and surgical procedures and ELISAs were performed to evaluate thrombus development and tissue factor (TF) activity. A total of 198 patients were found to be positive for IgA anti-β2 GPI isotype, and 57 patients were positive exclusively for IgA anti-β2 GPI antibodies. Of these, 13 of 23 patients (56.5%) in the LUMINA cohort, 17 of 17 patients (100%) in the Hopkins cohort, and 10 of 17 patients (58.9%) referred to APLS had at least one APS-related clinical manifestation. Fifty-four percent of all the IgA anti-β2 GPI-positive serum samples reacted with domain IV/V of anti-β2 GPI, and 77% of those had clinical features of APS. Isolated IgA anti-β2 GPI positivity was associated with an increased risk of arterial thrombosis (P < 0.001), venous thrombosis (P = 0.015), and all thrombosis (P < 0.001). The association between isolated IgA anti-β2 GPI and arterial thrombosis (P = 0.0003) and all thrombosis (P = 0.0003) remained significant after adjusting for other risk factors for thrombosis. In vivo mouse studies demonstrated that IgA anti-β2 GPI antibodies induced significantly larger thrombi and higher TF levels compared to controls. Isolated IgA anti-β2 GPI-positive titers may identify additional patients with clinical features of APS. Testing for these antibodies when other antiphospholipid tests are negative and APS is suspected is recommended. IgA anti-β2 GPI antibodies directed to domain IV/V of β2 GPI represent an important subgroup of clinically relevant antiphospholipids.

Heterogeneity between Diagnostic Tests for IgA anti-Beta2 Glycoprotein I: Explaining the Controversy in Studies of Association with Vascular Pathology

IgA antibeta 2 Glycoprotein I (β2GPI) antibodies test can identify some patients with antiphospholipid syndrome (APS) that are negative for other isotypes. Controversy exists because some studies have reported a strong association of these antibodies with vascular disease, while others have not confirmed this observation. Our hypothesis is that these contradictory results may be due to differences among commercial diagnostic kits. To answer this question, we have compared the results obtained with several of the most commonly used commercial IgA anti β2GPI antibodies (aβ2GPI) diagnostic assays on specimens from individuals suspected of having APS. Sera from 69 patients (37 positive and 32 negative for IgA aβ2GPI) were analyzed with seven different commercial ELISA kits for IgA aβ2GPI, following instructions and cutoffs provided by the manufacturer. Our results showed important differences in the sensitivity and specificity of the different assays. Two of the seven kits tested had a sensitivity level below 65% for IgA aβ2GPI, and three showed levels of specificity lower than 80%. Some commercial kits to detect IgA aβ2GPI are suboptimal. Variability between kits may account for the discrepancy in results obtained and for the lack of consensus concerning their clinical significance. It is important that the scientific community work to standardize assay performance so that the true clinical significance of this important clinical marker can be clearly established.

AB0726 The utility of IGA antiphospholipid antibodies for prediction of thrombotic complications in secondary antiphospholipid syndrome

BackgroundAt present we use modified classification criteria (Sydney 2006) for a diagnosis of antiphospholipid syndrome (APS) (1). These criteria do not include IgA class anticardiolipin (aCL) or antiβ2-glycoprotein I (antiβ2GPI)...

Antiphospholipid antibodies and atherosclerosis: Insights from Rheumatoid arthritis – A five-year follow-up study

Objectives Life expectancy in rheumatoid arthritis (RA) patients is reduced by 3–10 years, probably due to cerebrovascular and cardiovascular diseases associated with atherosclerosis. In the present study, we wanted to verify if previously reported IgA anti-beta 2-glycoprotein I (2GPI) antibodies possibly represented an independent risk factor for atherosclerosis in RA patients during a longer period of follow up. Methods The follow-up study (after 5.5 years) comprised all initially included patients and controls (premenopausal women, non-diabetic, normotensive at the start of the study), except for two RA patients (one died and one not available). The same clinical, laboratory and ultrasound assessments were performed. Results Patients and controls were divided into three categories: Intima-media thickness (IMT) progressors, plaque progressors, IMT and plaque progressors. In controls, 55% represented IMT progressors and 5% IMT and plaque progressors. No statistically significant differences were detected comparing the progressors with delta (Δ = difference between follow-up and baseline study for each group in a time span of 5.5 years) LDL cholesterol, homocysteine and IgA anti-β2GPI. In patients, there were 48.5% IMT progressors, 5.8% plaque progressors and 19.1% IMT and plaque progressors. The progression was statistically significant associated with the levels of Δ homocysteine and Δ apolipoprotein B but not with LDL cholesterol and IgA anti-β2GPI. Conclusions The follow-up study showed advanced atherosclerosis in RA patients compared to sex and age matched controls. However, we were not able to confirm our initial impression that IgA anti-β2GPI might represent an independent risk factor for atherosclerosis.

Antiphospholipid antibodies in HIV-positive patients

Antiphospholipid (aPL) antibodies classically have been associated with thrombotic phenomena and abortion in patients with autoimmune diseases. The objective of the present work was to evaluate the frequency of such antibodies in patients infected with HIV and study its association with the presence of clinical manifestations of antiphospholipid syndrome (APS). Using a transversal study, a population of patients diagnosed with HIV, identified through an enzyme-linked immunosorbent assay (ELISA) test and confirmed by Western blotting, aged above 17 years old, was investigated. Through a standard questionnaire, the presence of APS manifestations was investigated, as well as the frequency of rheumatic manifestations. Antibodies against beta2 glycoprotein I (anti-beta2 GPI) and anticardiolipin (aCL) IgA, IgG, and IgM were investigated by the ELISA method using commercial kits (QUANTA Lite, INOVA Diagnostics). Ninety patients were studied, 47 (52.2%) male and 43 (47.8%) female. Clinical manifestations of APS were detected in 12 patients (13.3%) of the studied population, whereas arthralgia was the most common rheumatic manifestation (38.9%). Of the 90 patients, 40 (44.4%) were reactive for at least one type of aPL antibody (aCL and/or anti-beta2 GPI). The frequency of aCL was 17.8%, from which 15 (16.7%) had aCL IgG, 3 (3.3%) IgM, and 1 (1.1%) IgA. The frequency of the anti-beta2 GPI antibody was 33.3%, from which 29 (32.2%) were positive for isotype IgA, 4 (4.4%) isotype IgM, and 1 (1.1%) isotype IgG. No association was observed between immunoreactivity for aPL antibodies in general or each isotype in particular and the presence of APS manifestation. In the present study, it was possible to observe a relatively high frequency of aPL antibodies, particularly for isotype IgA anti-beta2 GPI in HIV. However, there was no association to APS manifestations, suggesting that such antibodies had no etiopathogenic role in these complications in patients with such retroviral infection.

Patients with atherosclerotic syndrome, negative in anti-cardiolipin assays, make IgA autoantibodies that preferentially target domain 4 of β 2-GPI

Autoantibodies targeting β 2-glycoprotein l (β 2-GPI), a component of the atherosclerotic plaque, are commonly found in patients with acute ischemic syndromes. Serum samples from APS (antiphospholipid syndrome) patients and from cardiovascular patients exhibiting acute atherosclerotic syndromes were analyzed for IgG and IgA antibodies in both anti-β 2-GPI and anticardiolipin (aCL) ELISA assays. All of the APS samples used here were positive in both assays. Serum samples from 382 atherosclerosis patients were also analyzed for IgG and IgA antibodies in the same assays. In sharp contrast to the APS samples, we found that only 1% of the samples from atherosclerosis patients were positive for IgA aCL, and 1.6% positive for IgG aCL, whereas 35.6% were positive for IgA anti-β 2-GPI and only 1.6% for IgG anti-β 2-GPI. The antigenic specificity of 29 serum samples from atherosclerosis patients was evaluated. Six different recombinant domain-deleted mutants (DM) of human β 2-GPI and full-length human β 2-GPI (wild-type) were used in competitive inhibition assays to inhibit the autoantibodies from binding in the anti-β 2-GPI ELISA assays. Domain-deleted mutants D—345 and D---45 inhibited the binding in the IgA anti-β 2-GPI assay, suggesting that these autoantibodies recognize domain 4 of the β 2-GPI molecule. These results clearly show that IgA anti-β 2-GPI autoantibodies from atherosclerotic patients are distinct from IgA autoantibodies found in APS samples.

Standardized measurement of major immunoglobulin class (IgG, IgA, and IgM) antibodies to β2glycoprotein I in patients with antiphospholipid syndrome

Patients with the antiphospholipid syndrome (APS) have autoantibodies directed against epitopes on beta2 glycoprotein I (beta2GPI). We describe herein the performance characteristics of standardized enzyme-linked immunosorbent assays (ELISAs) for anti-beta2GPI of the three major immunoglobulin classes: IgG, IgA, and IgM. All three assays generated highly linear standard curves (5 points, r > or = 0.993 for each); precision was excellent both intra-assay and run-to-run, with coefficients of variation (CV) ranging from 2.3% to 6.6%. Values for IgG anti-beta2GPI correlated strongly with those obtained by an earlier method (r = 0.80, P< 0.0001). A study group consisting of 203 healthy subjects was used to generate percentile-based reference intervals for all three classes of anti-beta2GPI. APS subjects' anti-beta2GPI were found to differ significantly (P <0.0001 for each) from those of the control population. All three assays correlated well with beta2GPI-dependent anticardiolipin antibody (aCL) measurements; IgG: r = 0.94 (P <0.0001), for IgA: r = 0.82 (P<0.001) and for IgM: r = 0.84 (P<0.0001). We suggest that these ELISAs may provide valuable standardized measurements of IgG, IgA, and IgM anti-P2GPI.