The end of the laboratory developed test as we know it? Recommendations from a national multidisciplinary taskforce of laboratory specialists on the interpretation of the IVDR and its complications.

The in vitro diagnostic medical devices regulation (IVDR) will take effect in May 2022. This regulation has a large impact on both the manufacturers of in vitro diagnostic medical devices (IVD) and clinical laboratories. For clinical laboratories, the IVDR poses restrictions on the use of laboratory developed tests (LDTs). To provide a uniform interpretation of the IVDR for colleagues in clinical practice, the IVDR Task Force was created by the scientific societies of laboratory specialties in the Netherlands. A guidance document with explanations and interpretations of relevant passages of the IVDR was drafted to help laboratories prepare for the impact of this new legislation. Feedback from interested parties and stakeholders was collected and used to further improve the document. Here we would like to present our approach to our European colleagues and inform them about the impact of the IVDR and, importantly we would like to present potentially useful approaches to fulfill the requirements of the IVDR for LDTs.

Multicentre comparison of a diagnostic assay: aquaporin-4 antibodies in neuromyelitis optica.

OBJECTIVE: Antibodies to cell surface central nervous system proteins help to diagnose conditions which often respond to immunotherapies. The assessment of antibody assays needs to reflect their clinical utility. We report the results of a multicentre study of aquaporin (AQP) 4 antibody (AQP4-Ab) assays in neuromyelitis optica spectrum disorders (NMOSD). METHODS: Coded samples from patients with neuromyelitis optica (NMO) or NMOSD (101) and controls (92) were tested at 15 European diagnostic centres using 21 assays including live (n=3) or fixed cell-based assays (n=10), flow cytometry (n=4), immunohistochemistry (n=3) and ELISA (n=1). RESULTS: Results of tests on 92 controls identified 12assays as highly specific (0-1 false-positive results). 32 samples from 50 (64%) NMO sera and 34 from 51 (67%) NMOSD sera were positive on at least two of the 12 highly specific assays, leaving 35 patients with seronegative NMO/spectrum disorder (SD). On the basis of a combination of clinical phenotype and the highly specific assays, 66 AQP4-Ab seropositive samples were used to establish the sensitivities (51.5-100%) of all 21 assays. The specificities (85.8-100%) were based on 92 control samples and 35 seronegative NMO/SD patient samples. CONCLUSIONS: The cell-based assays were most sensitive and specific overall, but immunohistochemistry or flow cytometry could be equally accurate in specialist centres. Since patients with AQP4-Ab negative NMO/SD require different management, the use of both appropriate control samples and defined seronegative NMOSD samples is essential to evaluate these assays in a clinically meaningful way. The process described here can be applied to the evaluation of other antibody assays in the newly evolving field of autoimmune neurology.

Therapeutic drug monitoring of infliximab: performance evaluation of three commercial ELISA kits.

BACKGROUND: Therapeutic drug monitoring (TDM) of infliximab (IFX, Remicade®) can aid to optimize therapy efficacy. Many assays are available for this purpose. However, a reference standard is lacking. Therefore, we evaluated the analytical performance, agreement and clinically relevant differences of three commercially available IFX ELISA kits on an automated processing system. METHODS: The kits of Theradiag (Lisa Tracker Infliximab), Progenika (Promonitor IFX) and apDia (Infliximab ELISA) were implemented on an automated processing system. Imprecision was determined by triplicate measurements of patient samples on five days. Agreement was evaluated by analysis of 30 patient samples and four spiked samples by the selected ELISA kits and the in-house IFX ELISA of Sanquin Diagnostics (Amsterdam, The Netherlands). Therapeutic consequences were evaluated by dividing patients into four treatment groups using cut-off levels of 1, 3 and 7 µg/mL and determining assay concordance. RESULTS: Within-run and between-run imprecision were acceptable (≤12% and ≤17%, respectively) within the quantification range of the selected ELISA kits. The apDia assay had the best precision and agreement to target values. Statistically significant differences were found between all assays except between Sanquin Diagnostics and the Lisa Tracker assay. The Promonitor assay measured the lowest IFX concentrations, the apDia assay the highest. When patients were classified in four treatment categories, 70% concordance was achieved. CONCLUSIONS: Although all assays are suitable for TDM, significant differences were observed in both imprecision and agreement. Therapeutic consequences were acceptable when patients were divided in treatment categories, but this could be improved by assay standardization.

Anti-MOG antibodies plead against MS diagnosis in an Acquired Demyelinating Syndromes cohort.

BACKGROUND: Acquired demyelinating syndromes (ADS) in children are a group of distinct first immune-mediated demyelinating events of the central nervous system (CNS). Predictive biomarkers for future diagnosis are lacking. A putative target antigen is myelin oligodendrocyte glycoprotein (MOG). We analyzed the presence of MOG antibodies in a cohort of ADS patients in The Netherlands. METHODS: Using a cell-based assay, we analyzed 117 children with ADS from a nationwide cohort, whom were divided into five groups: optic neuritis (ON; n = 20), transverse myelitis (TM; n = 7), other monofocal ADS (n = 22), polyfocal ADS without encephalopathy (n = 44) and polyfocal ADS with encephalopathy (n = 24). Additionally, we tested children with other neurological diseases (OND; n = 13), healthy children (n = 31) and adult polyfocal ADS plus encephalopathy (ADEM) patients (n = 29). RESULTS: We found that 21 of the 117 children with ADS tested anti-MOG seropositive (18%). The group of patients with ADEM had the highest prevalence of anti-MOG seropositivity (42% versus 18% in the non-encephalopathic polyfocal ADS patients). Although 47 ADS children had a final diagnosis of multiple sclerosis (MS), in only one of them were MOG antibodies detected (2%), with only borderline positivity. Only 1 out of the 29 adult ADEM patients tested anti-MOG seropositive. CONCLUSIONS: MOG antibodies are strongly skewed towards ADS children that present with an ADEM-like disease onset. The presence of such antibodies pleads against a future diagnosis of MS.

Anti-SSA antibodies are present in immunoglobulin preparations.

BACKGROUND: Anti-SSA autoantibodies are among the most frequently detected autoantibodies and have traditionally been associated with Sjögren's syndrome (SjS) and systemic lupus erythematosus. The unexpected finding of anti-SSA antibodies in a patient with common variable immunodeficiency disorder (CVID) treated with intravenous immunoglobulin (IVIG), who developed discoid lupus erythematosus, prompted us to investigate the presence of anti-SSA antibodies in IVIG preparations. Since anti-SSA antibodies may be present in apparently healthy individuals without overt autoimmune features, IVIG preparations may also contain anti-SSA antibodies. STUDY DESIGN AND METHODS: IVIG consists of polyclonal immunoglobulin G isolated from the plasma of more than 1000 blood donors. Several IVIG batches from different suppliers and serum samples of patients receiving these IVIG products were tested for the presence of anti-nuclear antibodies (ANAs) and extractable nuclear antibodies (ENAs). In addition, we tested several plasma pools for the presence of anti-SSA and subsequent serum samples of individual donors. RESULTS: Several CVID-patients receiving IVIG tested positive for ANA and anti-SSA. The IVIG products administered also contained clearly detectable concentrations of these antibodies. The frequency of apparently healthy blood donors with anti-SSA positivity was 0.69% and one of 1894 donors (0.05%) showed a very high titer of anti-SSA of more than 10,000 U/mL. CONCLUSION: Anti-SSA is present in IVIG products and in blood donors without clinical symptoms. IVIG replacement can interfere with ANA and ENA serology by passive transfer of autoantibodies. We hypothesize that such autoantibodies may be causally related to disease manifestations in some recipients.

Anti-carbamylated protein (anti-CarP) antibodies precede the onset of rheumatoid arthritis.

OBJECTIVE: The presence of anti-citrullinated protein antibodies (ACPA) and IgM-rheumatoid factor (IgM-RF) years before the clinical diagnosis of rheumatoid arthritis (RA) suggests they are possibly involved in the pathogenic process underlying RA. In this study, we analysed whether anti-carbamylated protein (anti-CarP) antibodies, a novel autoantibody system against carbamylated proteins, can also be detected in healthy individuals before they developed RA. METHODS: Multiple sera from asymptomatic blood donors prior to the onset of their RA symptoms and sera from age-matched and sex-matched controls were tested for the presence of antibodies directed against carbamylated-fetal calf serum (Ca-FCS), carbamylated-fibrinogen (Ca-Fib), cyclic citrullinated-peptide 2 and IgM-RF. RESULTS: Anti-Ca-FCS and anti-Ca-Fib antibodies were each present in 27% and 38% of the last serum samples of blood donors prior to the diagnosis of RA. Both anti-Ca-FCS and anti-Ca-Fib antibodies could be detected many years before the onset of RA. Anti-CarP antibodies as well as ACPA are, on average, detected earlier than IgM-RF. CONCLUSIONS: In addition to ACPA and IgM-RF, also the newly identified anti-CarP antibodies appear many years before the diagnosis of RA.

Rising incidence and prevalence of primary biliary cirrhosis: a large population-based study.

BACKGROUND & AIMS: Large population-based studies are much needed to accurately establish the epidemiology of primary biliary cirrhosis (PBC). We aimed to collect all PBC patients in a geographically defined area to evaluate the epidemiology of PBC and examine the possible association of PBC with smoking, age at menarche, age at first pregnancy and number of pregnancies. METHODS: All PBC patients between 2000 and 2008 were identified in a geographically defined area of the Netherlands, comprising 50% of the Dutch population. Four independent hospital databases were searched in 44 hospitals. Medical records were reviewed on site verifying diagnosis and for collection of clinical data. Age- and gender matched controls were recruited from the outpatient clinics of four participating hospitals. Patients and controls were asked to fill out a questionnaire regarding family history, previous and current smoking behaviour and fertility status. RESULTS: Nine hundred and ninety-two PBC patients fulfilled all inclusion criteria, resulting in a mean incidence of 1.1 per 100 000; 0.3 in men and 1.9 in women. On January 1st 2008 the point prevalence was 13.2 per 100 000 inhabitants. Incidence and prevalence rates were increasing over time (P < 0.001). No geographical differences in disease distribution were observed. Smoking behaviour, age at menarche, age at first pregnancy, gravidity and number of children were not significantly different between cases and controls. CONCLUSION: Incidence and prevalence rates of PBC are increasing over time. PBC was not found to be associated with smoking, age at menarche, age at first pregnancy or number of pregnancies.

Monoclonal anti-citrullinated protein antibodies selected on citrullinated fibrinogen have distinct targets with different cross-reactivity patterns.

OBJECTIVE: ACPAs are thought to play a pathogenic role in RA. Because of their polyclonal nature it is difficult to study characteristics of ACPAs such as cross-reactivity or affinity. This study aimed to analyse the ACPA response at clonal level. METHODS: Citrullinated fibrinogen-specific B cells were isolated from blood derived from an RA patient by fluorescent automated cell sorting (FACS). Antigen specificity was verified by ELISA of culture supernatant. RNA of antigen-specific B cells was isolated and VH and VL chains were cloned and subsequently expressed as IgG1 antibodies. RESULTS: Two human recombinant antibodies were obtained that bind to citrullinated fibrinogen peptide (cFib). Both monoclonal antibodies originate from different naive B cells, undergo extensive somatic hypermutation and bind to cFib (but not to Fib) with moderate avidity. Furthermore, they show distinct cross-reactivity patterns towards other citrullinated peptides, suggesting that both antibodies have different primary targets. CONCLUSION: Together these data suggest that ACPAs are formed by antigen-driven maturation, and that multiple citrullinated antigens are involved in activating the B-cell response.

Maggot excretions affect the human complement system.

The complement system plays an important role in the activation of the inflammatory response to injury, although inappropriate complement activation (CA) can lead to severe tissue damage. Maggot therapy is successfully used to treat infected wounds. In this study, we hypothesized that maggot excretions/secretions influence CA in order to modulate the host's inflammatory response. Therefore, the effect of maggot excretions on CA was investigated in preoperatively and postoperatively obtained sera from patients. Our results show that maggot excretions reduce CA in healthy and postoperatively immune-activated human sera up to 99.9%, via all pathways. Maggot excretions do not specifically initiate or inhibit CA, but break down complement proteins C3 and C4 in a cation-independent manner and this effect proves to be temperature tolerant. This study indicates a CA-reducing substrate that is already successfully used in clinical practice and may explain part of the improved wound healing caused by maggot therapy. Furthermore, the complement activation-reducing substance present in maggot excretions could provide a novel treatment modality for several diseases, resulting from an (over)active complement system.

Development of the anti-citrullinated protein antibody repertoire prior to the onset of rheumatoid arthritis.

OBJECTIVE: To examine how anti-citrullinated protein antibody (ACPA) epitope spreading takes place prior to the onset of clinical rheumatoid arthritis (RA), and to analyze the pattern of autoantigen reactivity at the beginning of the immune response. METHODS: Multiple consecutive serum samples from 79 RA patients who had donated blood before disease onset were available for analysis. Fifty-three patients tested positive for ACPAs prior to the onset of clinical RA. For these patients, a median of 6 (interquartile range 4-9) sequential pre-RA serum samples obtained 1-2 years apart were tested. Reactivity to 5 distinct citrullinated peptides was measured by enzyme-linked immunosorbent assay. Two peptides were derived from fibrinogen, 1 from vimentin, 1 from α-enolase, and 1 from filaggrin. RESULTS: In 25 of 53 ACPA-positive patients, seroconversion from ACPA absence to ACPA presence was observed. In 72% of these patients, the immune response started with reactivity to 1 peptide, without preference for a particular peptide. The number of peptides recognized increased over time, without a dominant epitope-spreading pattern. ACPAs appeared in low levels several years prior to the diagnosis of RA. Antibody titers increased markedly ∼2-4 years before diagnosis. CONCLUSION: Our findings indicate that ACPA epitope spreading occurs over several years prior to the onset of clinical RA. The initial autoimmune response is mostly directed toward only 1 autoantigen, but this is not always the same antigen. The marked increase in ACPA titers a few years prior to the diagnosis of RA suggests a second stage in disease development, which might be due to a variety of factors.

The extent of the anti-citrullinated protein antibody repertoire is associated with arthritis development in patients with seropositive arthralgia.

OBJECTIVES: To determine the fine specificity of anti-citrullinated protein antibodies (ACPA) in the early phase of arthritis development, the ACPA repertoire in arthralgia patients and the association with arthritis development were studied. METHODS: A total of 244 patients with arthralgia positive for anti-cyclic citrullinated peptide antibodies (aCCPs) and/or IgM rheumatoid factor (IgM-RF), without arthritis were included. Development of arthritis was defined as presence of one or more swollen joints at clinical examination during follow-up. Sera were tested at baseline for reactivity to five citrullinated peptides derived from fibrinogen (three), vimentin (one) and α-enolase (one) and five corresponding arginine peptides in an ELISA. RESULTS: In all, 69 patients (28%) developed arthritis in a median of 3 joints after a median follow-up of 11 (IQR 5-20) months. Reactivity to each peptide was significantly associated with arthritis development (p<0.001). The ACPA repertoire did not differ between patients who did or did not develop arthritis. Among aCCP-positive patients, patients recognising two or more additional citrullinated peptides developed arthritis more often (p=0.04). The number of recognised peptides was positively associated with the aCCP level (p<0.001). Crossreactivity between different peptides was minimal. CONCLUSIONS: Arthritis development is not associated with recognition of a specific citrullinated peptide once joint complaints are present. The ACPA repertoire in some patients with arthralgia is expanded. High aCCP levels are associated with a qualitatively broad ACPA repertoire. Patients with an extended ACPA repertoire have a higher risk of developing arthritis.

Antibodies against aquaporin-4 in neuromyelitis optica: distinction between recurrent and monophasic patients.

The detection of antibodies against aquaporin-4 (AQP4) has improved the diagnosis of neuromyelitis optica (NMO). We evaluated a recently established cell-based anti-AQP4 assay in 273 patients with inflammatory CNS demyelination. The assay had a specificity of 99% and a sensitivity of 56% to detect all NMO patients and of 74% to detect the recurrent NMO patients, similar to the initial studies reported. AQP4 antibodies were absent in monophasic NMO patients, while samples in recurrent cases remained positive during follow-up. We conclude that the pathogenesis of monophasic NMO may be different from that of relapsing NMO.

High-throughput analysis of the C4 polymorphism by a combination of MLPA and isotype-specific ELISA's.

Complement factor C4 exists as two main isotypes, C4A and C4B, with different functional properties and encoded by two separate genes. In addition, C4A and C4B genes can occur in multiple copies and may carry a retroviral HERV-K(C4) insertion in intron 9. To study association of C4 polymorphism with disease, accurate genotyping and phenotyping is important. However, current techniques are very laborious and not suitable to study large patient groups. Therefore, we aimed to develop novel assays for C4 geno- and phenotyping, to make high throughput possible. To study C4 gene copy number variation, a novel Multiplex Ligation-dependent Probe Amplification (MLPA) assay was set up with three synthetic probe parts. C4A and C4B protein levels were measured by isotype-specific ELISA's. The relationship between C4 genotype with C4A and C4B serum concentrations was examined in 104 healthy lab workers and 66 children with meningococcal disease. As expected, a strong positive correlation was found between C4A and C4B gene copy number and serum levels of total C4, C4A and C4B. In the healthy controls, 95.3% of C4A genes and 53.7% of C4B genes carried the HERV-K(C4) insertion. Presence of HERV-K(C4) resulted in less C4B protein expression, while there was no effect on total C4 levels. In the meningitis patients, no increased incidence of hetero- or homozygous deficiency of either C4A or C4B was found. In conclusion, the combination of MLPA and ELISA is very suitable to study the geno- and phenotype of complement C4 in large patient groups.

Biomarkers of Checkpoint Inhibitor Induced Immune-Related Adverse Events-A Comprehensive Review.

Immune checkpoint inhibitors (ICIs) have substantially improved the prognosis of patients with different types of cancer. Through blockade of cytotoxic T-lymphocyte antigen 4 (CTLA-4) and programmed cell death protein 1 (PD-1), negative feedback mechanisms of the immune system are inhibited, potentially resulting in very durable anti-tumor responses. Despite their promise, ICIs can also elicit auto-immune toxicities. These immune-related adverse events (irAEs) can be severe and sometimes even fatal. Therefore, being able to predict severe irAEs in patients would be of added value in clinical decision making. A search was performed using "adverse events", "immune checkpoint inhibitor", "biomarker", and synonyms in PubMed, yielding 3580 search results. After screening title and abstract on the relevance to the review question, statistical significance of reported potential biomarkers, and evaluation of the remaining full papers, 35 articles were included. Five additional reports were obtained by means of citations and by using the similar article function on PubMed. The current knowledge is presented in comprehensive tables summarizing blood-based, immunogenetic and microbial biomarkers predicting irAEs prior to and during ICI therapy. Until now, no single biomarker has proven to be sufficiently predictive for irAE development. Recommendations for further research on this topic are presented.

Autoantibodies in idiopathic inflammatory myopathies: Clinical associations and laboratory evaluation by mono- and multispecific immunoassays.

Idiopathic inflammatory myopathies (IIM) are a group of diseases characterized by immune-mediated muscular lesions that may be associated with extra-muscular manifestations involving skin, lungs, heart or joints. Four main groups of IIM can be distinguished: dermatomyositis (DM), overlap myositis including mainly anti-synthetase syndrome (ASS), immune mediated necrotizing myopathy (IMNM), and inclusion body myositis (IBM). Myositis-specific autoantibodies (MSA) are increasingly recognized as valuable tools for diagnosis, classification and prognosis of IIM. For example, ASS is associated with anti-aminoacyl tRNA synthetase antibodies (anti-Jo-1, PL-7, PL-12, …), IMNM with anti-SRP and anti-HMGCR; IBM with anti-cytosolic 5'nucleotidase 1A (cN1A), and DM with anti-Mi-2, anti-MDA-5, anti-TIF-1γ, anti-NXP-2 and anti-SAE. Moreover, anti-MDA-5 is associated with amyopathic myositis and interstitial lung disease and anti-TIF-1γ and anti-NXP-2 with juvenile DM as well as malignancy in patients >40 years. Most MSA have initially been discovered by immunoprecipitation. In routine laboratories, however, MSA are screened for by indirect immunofluorescence and identified by (automated) monospecific immunoassays or by multispecific immunoassays (mainly line/dot immunoassays). Validation of these (multispecific) assays is a challenge as the antibodies are rare and the assays diverse. In this review, we give an overview of the (clinical) performance characteristics of monospecific assays as well as of multispecific assays for detection of MSA. Although most assays are clinically useful, there are differences between techniques and between manufacturers. We discuss that efforts are needed to harmonize and standardize detection of MSA.

Frequencies and clinical associations of myositis-related antibodies in The Netherlands: A one-year survey of all Dutch patients.

Idiopathic inflammatory myopathies (IIM) are a heterogeneous group of connective tissue diseases, collectively known as myositis. Diagnosis of IIM is challenging while timely recognition of an IIM is of utter importance considering treatment options and otherwise irreversible (severe) long-term clinical complications. With the EULAR/ACR classification criteria (2017) considerable advancement has been made in the diagnostic workup of IIM. While these criteria take into account clinical parameters as well as presence of one autoantibody, anti-Jo-1, several autoantibodies are associated with IIM and are currently evaluated to be incorporated into classification criteria. As individual antibodies occur at low frequency, the development of line blots allowing multiplex antibody analysis has improved laboratory diagnostics for IIM. The Euroline myositis line-blot assay (Euroimmun) allows screening and semi-quantitative measurement for 15 autoantibodies, i.e. myositis specific antibodies (MSA) to SRP, EJ, OJ, Mi-2α, Mi-2ß, TIF1-γ, MDA5, NXP2, SAE1, PL-12, PL-7, Jo-1 and myositis associated antibodies (MAA) to Ku, PM/Scl-75 and PM/Scl-100. To evaluate the clinical significance of detection and levels of these autoantibodies in the Netherlands, a retrospective analysis of all Dutch requests for extended myositis screening within a 1 year period was performed. A total of 187 IIM patients and 632 non-IIM patients were included. We conclude that frequencies of MSA and MAA observed in IIM patients in a routine diagnostic setting are comparable to cohort-based studies. Weak positive antibody levels show less diagnostic accuracy compared to positive antibody levels, except for anti-NXP2. Known associations between antibodies and skin involvement (anti-MDA5, anti-TIF1-γ), lung involvement (anti-Jo-1), and malignancy (anti-TIF1-γ) were confirmed in our IIM study population. The availability of multiplex antibody analyses will facilitate inclusion of additional autoantibodies in clinical myositis guidelines and help to accelerate diagnosing IMM with rare but specific antibodies.

High incidence of type 1 diabetes mellitus during or shortly after treatment with pegylated interferon alpha for chronic hepatitis C virus infection.

BACKGROUND: Development of diabetes mellitus (DM) during or shortly after treatment with interferon alpha (IFN-alpha) in patients with chronic hepatitis C virus (HCV) infection has been reported sporadically. We prospectively screened for DM during and after IFN-alpha therapy for chronic HCV infection. METHODS: Blood glucose levels of patients with chronic HCV infection were routinely assessed at all outpatient visits during and after treatment with pegylated-IFN-alpha (Peg-IFN-alpha) and ribavirin (Riba). RESULTS: Between December 2002 and October 2005, 189 non-diabetic patients were treated with Peg-IFN-alpha/Riba, of whom five developed type 1 DM (2.6%), three type 2 DM (1.6%) and one an indeterminate type of DM. Classical symptoms of DM were present in three patients who developed DM shortly after cessation of Peg-IFN-alpha/Riba. In the other patients, symptoms of DM were either indistinguishable from side effects caused by Peg-IFN-alpha/Riba or absent. CONCLUSION: Our study showed a high incidence of type 1 DM during Peg-IFN-alpha/Riba therapy for chronic HCV infection. Symptoms of DM may be absent or mistaken for Peg-IFN-alpha/Riba-associated side effects. To diagnose DM without delay, we propose routine assessment of blood glucose at all outpatient visits during and after Peg-IFN-alpha/Riba treatment in chronic HCV patients.

Comparisons of Serum Infliximab and Antibodies-to-Infliximab Tests Used in Inflammatory Bowel Disease Clinical Trials of Remicade®.

Monitoring infliximab (IFX) concentrations and antibodies-to-IFX (ATI) titers during inflammatory bowel disease treatment may allow more informed decisions in assessing exposure/response and determining appropriate dosing. To aid in interpreting results from different commercial tests in the context of Janssen's published Remicade® results, the reliability of Janssen's IFX and ATI assays was compared with commercial assays from KU Leuven, Sanquin, Dynacare, and LabCorp. Test results were independently reported to Janssen. All assays were tested for specificity, selectivity, and precision. ATI assays were evaluated for sensitivity, drug interference, and potential interference of tumor necrosis factor-alpha (TNF-α). IFX assays were specific, accurate, and reproducible. Intra-class correlation of Janssen IFX assay results with those from KU Leuven, Sanquin, Dynacare, and LabCorp were 0.960, 0.895, 0.931, and 0.971, respectively. ATI titers >10 interfered with IFX assessment in all IFX assays, whereas TNF-α (≤50 ng/mL) did not interfere with IFX detection in any assay. ATI assays specifically and reproducibly detected ATI. Janssen, Sanquin, and LabCorp ATI methods were more resistant to IFX interference than Dynacare and KU Leuven, which were affected by IFX concentrations at ≥2 µg/mL. TNF-α (<5 ng/mL) did not interfere with ATI detection. Strong agreement was observed between Janssen's IFX and ATI assays and the diagnostic service provider assays. Our study results indicate that all four commercially available assays are suitable for therapeutic drug monitoring of IFX. The substantial agreement reported here between the comparator assays and the Janssen drug-tolerant assay provides support to clinicians in their use of these commercial assays, and for understanding their patients' IFX and ATI results relative to published data from clinical studies of Remicade.