Assessing serum anti-nuclear antibodies HEp-2 patterns in synucleinopathies.

This study investigates the presence of antinuclear antibodies (ANA) in three primary synucleinopathies - Parkinson's disease (PD), multiple system atrophy (MSA), and dementia with Lewy bodies (DLB), compared to healthy controls. Autoinflammatory disorders typically involve the immune system mistakenly attacking the body's own cells and start producing ANA. There is an increasing body of evidence that immune-mediated inflammation is a pathological feature linked to synucleinopathies. To investigate whether this could be autoimmune mediated we analyzed for ANA in the plasma of 25 MSA, 25 PD, and 17 DLB patients, along with 25 healthy controls, using the ANA HEp-2 indirect immunofluorescence antibody assay (ANA HEp-2 IFA). Contrary to initial expectations, results showed ANA HEp-2 positivity in 12% of PD, 8% of MSA patients, 18% of DLB patients, and 17% of healthy controls, indicating no increased prevalence of ANA in synucleinopathies compared to age-matched healthy individuals. Various ANA HEp-2 patterns were identified, but no specific pattern was associated with individual synucleinopathies. We conclude hereby that synucleinopathies are not associated with detectable presence of ANA in plasma.

Validation of an indirect ELISA assay for assessment of autoantibodies against full-length TRIM21 and its individual domains.

Anti-SSA-autoantibodies are common in patients with rheumatologic disease, especially Sjögren's syndrome, systemic lupus erythematosus and rheumatoid arthritis. They consist of both autoantibodies towards Ro60 and Ro52, the latter also known as TRIM21. TRIM21 is an intracellular protein consisting of four domains; PRY/SPRY, Coiled-Coil, B-box and RING. The aim of this study was to establish an indirect ELISA detecting autoantibodies towards both the full-length TRIM21 protein and its four domains. We expressed the five constructs, created, and validated indirect ELISA protocols for each target using plasma from anti-SSA positive patients and healthy controls. Our findings were validated to the clinically used standards. We measured significantly higher levels of autoantibodies towards our full-length TRIM21, and the PRY/SPRY, Coiled-Coil and RING domains in patients compared to healthy controls. No significant difference in the level of autoantibodies were detected against the B-box domain. Our setups had a signal to noise ratio in the range of 30 to 184, and an OD between 2 and 3. Readings did not decline using NaCl of 500 mM as wash, affirming the high binding affinity of the autoantibodies measured. Our protocols allow us to further study the different autoantibodies of anti-SSA positive patients. This creates the possibility to stratify our patients into subgroups regarding autoantibody profile and specific pheno- or endotype.

Comparison of 16 Serological SARS-CoV-2 Immunoassays in 16 Clinical Laboratories.

Serological assays for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are needed to support clinical diagnosis and epidemiological investigations. Recently, assays for large-scale detection of total antibodies (Ab), immunoglobulin G (IgG), and IgM against SARS-CoV-2 antigens have been developed, but there are limited data on the diagnostic accuracy of these assays. This study was a Danish national collaboration and evaluated 15 commercial and one in-house anti-SARS-CoV-2 assays in 16 laboratories. Sensitivity was evaluated using 150 samples from individuals with asymptomatic, mild, or moderate COVID-19, nonhospitalized or hospitalized, confirmed by nucleic acid amplification tests (NAAT); samples were collected 13 to 73 days either from symptom onset or from positive NAAT (patients without symptoms). Specificity and cross-reactivity were evaluated in samples collected prior to the SARS-CoV-2 epidemic from >586 blood donors and patients with autoimmune diseases, cytomegalovirus or Epstein-Barr virus infections, and acute viral infections. A specificity of ≥99% was achieved by all total-Ab and IgG assays except one, DiaSorin Liaison XL IgG (97.2%). Sensitivities in descending order were Wantai ELISA total Ab (96.7%), CUH-NOVO in-house ELISA total Ab (96.0%), Ortho Vitros total Ab (95.3%), YHLO iFlash IgG (94.0%), Ortho Vitros IgG (93.3%), Siemens Atellica total Ab (93.2%), Roche Elecsys total Ab (92.7%), Abbott Architect IgG (90.0%), Abbott Alinity IgG (median 88.0%), DiaSorin Liaison XL IgG (median 84.6%), Siemens Vista total Ab (81.0%), Euroimmun/ELISA IgG (78.0%), and Snibe Maglumi IgG (median 78.0%). However, confidence intervals overlapped for several assays. The IgM results were variable, with the Wantai IgM ELISA showing the highest sensitivity (82.7%) and specificity (99%). The rate of seropositivity increased with time from symptom onset and symptom severity.

No evidence of increased anti-M3 muscarinic acetylcholine receptor autoantibodies in SSA-positive connective tissue disease patients.

Anti-muscarinic type 3 receptor autoantibodies (M3R) and anti-SSA antibodies are both related to salivary secretion. The presence of M3R antibodies in Sjögren's syndrome is previously demonstrated; nevertheless, the relationship between the anti-SSA antibodies and M3R fragment antibodies, namely the N terminal, first, second, and third extracellular loops, remains to be elucidated. In this study, we analyzed the antibodies against the M3R epitopes in healthy controls and anti-SSA antibody-positive connective tissue disease patients through ELISA method. Antibodies against the first, second, and third extracellular loop (M3R211-230 ) were not increased in anti-SSA positive patients compared to healthy controls. Indeed, antibodies against the N terminal (M3R1-33 ) were found to be high in healthy controls. High levels of M3R1-33 in healthy controls are a novel original finding; further research is needed for the clinical significance. There is no significant difference between SSA-positive patients and healthy controls in terms of autoantibodies against the remainder of the linear M3R fragments.