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.

Formation and Glomerular Deposition of Immune Complexes in Mice Administered Human Antibodies: Evaluation of Dose, Frequency, and Biomarkers.

Administration of human protein-based drugs to animals often leads to formation of antidrug antibodies (ADAs) that may form circulating immune complexes (CICs) with the dosed protein. Circulating immune complexes can activate and bind complement (cCICs), and if large amount of CICs or cCICs is formed, the clearance mechanism potentially becomes saturated, which can lead to immune complex (IC) deposition and inflammation. To obtain a better understanding of the underlying factors, including the relationship between different dose regimes on IC formation and deposition and identification of possible biomarkers of IC deposition and IC-related pathological changes in kidneys, BALB/c and C57BL/6J mice were administered with human anti-tumor necrosis factor α (aTNFα, adalimumab) or a humanized anti-TNP (aTNP) antibody for 13 weeks. Particularly, ADA, CIC, cCIC formation, IC deposition, and glomerulonephritis were observed in C57BL/6J administered with aTNFα, whereas the immunologic response was minor in BALB/c mice administered with aTNFα and in BALB/c and C57BL/6J mice administered aTNP. Changing dose levels or increasing dosing frequency of aTNFα on top of an already-established CIC and cCIC response did not lead to substantial changes in CIC, cCIC formation, or IC deposition. Finally, no association between the presence of CICs or cCIC in plasma and glomerular IC deposition and/or glomerulonephritis was observed.

Formation and glomerular deposition of immune complexes in mice administered bovine serum albumin: Evaluation of dose, frequency, and biomarkers.

In preclinical toxicity studies, species-foreign proteins administered to animals frequently leads to formation of anti-drug antibodies (ADA). Such antibodies may form circulating immune complexes (CIC) with the administered protein. These CIC can activate the classical complement pathway, thereby forming complement-bound CIC (cCIC); if large of amounts of CIC or cCIC is formed, the clearance mechanism may become saturated which potentially leads to vascular immune complex (IC) deposition and inflammation. Limited information is available on the effect of different treatment related procedures as well as biomarkers of IC-related vascular disease. In order to explore the effect of different dose regimens on IC formation and deposition, and identification of possible biomarkers of IC deposition and IC-related pathological changes, C57BL/6J and BALB/c mice were dosed subcutaneously twice weekly with bovine serum albumin (BSA) for 13 weeks without adjuvant. After 6 and 13 weeks, CIC and cCIC were detected in plasma; after 13 weeks, IC deposition was detected in kidney glomeruli. In particular immunohistochemistry double-staining was shown to be useful for detection of IC deposition. Increasing dosing frequency or changing BSA dose level on top of an already established CIC and cCIC response did not cause changes in IC deposition, but CIC and cCIC concentrations tended to decrease with increased dose level, and increased cCIC formation was observed after more frequent dosing. The presence of CIC in plasma was associated with glomerular IC deposits in the dose regimen study; however, the use of CIC or cCIC as potential biomarkers for IC deposition and IC-related pathological changes, needs to be explored further.

Generic immune complex assay for detection of murine anti-drug-antibodies in complex with human IgG.

Rapid and versatile methods are needed for evaluation of immunogenicity in early safety studies. The present work presents a generic, simple and easy to use sandwich enzyme-linked immunosorbent assay for quasi-quantitative measurement of circulating immune complexes (CICs) formed by anti-drug antibodies (ADAs) in complex with human IgG in mouse plasma. The assay is suitable for evaluating the presence of in vivo formed CICs in mice exposed to human IgG antibodies independent of target and IgG subtype. The assay is established using commercially available antibodies, and calibrated using CIC mimics based on bis(sulfosuccinimidyl)suberate conjugated human and mouse IgG. The development and qualification process of the generic methodology is described and include acceptance criteria, stability, sensitivity, drug tolerance, spike recovery, precision and cut point determination. In order to demonstrate assay performance, its use is exemplified by quantifying CICs in mice administered with a fully human anti-TNF-α IgG1 antibody (adalimumab) or a humanized anti-trinitrophenol (TNP) IgG4 antibody. Results show a well-qualified reproducible assay set-up with adequate sensitivity, easy discrimination between positive and negatives and quasi-quantitative measurement of ADA-human IgG CICs in mice administered with each of two different human/humanized IgG antibodies.