Bridging ELISA Research Articles

Prokaryotic expression and characterization of the heterodimeric construction of ZnT8 and its application for autoantibodies detection in diabetes mellitus

BackgroundIn the present work we described the recombinant production and characterization of heterodimeric construction ZnT8-Arg-Trp325 fused to thioredoxin using a high-performance expression system such as Escherichia coli. In addition, we apply this novel recombinant antigen in a non-radiometric method, with high sensitivity, low operational complexity and lower costs.ResultsZnT8 was expressed in E. coli as a fusion protein with thioredoxin (TrxZnT8). After 3 h for induction, recombinant protein was obtained from the intracellular soluble fraction and from inclusion bodies and purified by affinity chromatography. The expression and purification steps, analyzed by SDS-PAGE and western blot, revealed a band compatible with TrxZnT8 expected theoretical molecular weight (≈ 36.8 kDa). The immunochemical ability of TrxZnT8 to compete with [35S]ZnT8 (synthesized with rabbit reticulocyte lysate system) was assessed qualitatively by incubating ZnT8A positive patient sera in the presence of 0.2–0.3 μM TrxZnT8. Results were expressed as standard deviation scores (SDs). All sera became virtually negative under antigen excess (19.26–1.29 for TrxZnT8). Also, radiometric quantitative competition assays with ZnT8A positive patient sera were performed by adding TrxZnT8 (37.0 pM–2.2 µM), using [35S]ZnT8. All dose–response curves showed similar protein concentration that caused 50% inhibition (14.9–0.15 nM for TrxZnT8). On the other hand, preincubated bridge ELISA for ZnT8A detection was developed. This assay showed 51.7% of sensitivity and 97.1% of specificity.ConclusionsIt was possible to obtain with high-yield purified heterodimeric construction of ZnT8 in E. coli and it was applied in cost-effective immunoassay for ZnT8A detection.

A simple and cost-effective assay for measuring anti-drug antibody in human patients treated with Adalimumab

It has been reported that 90% of the anti-drug antibody (ADA) to Adalimumab in human patients bound to the TNF-binding area, resulted in the annual loss of responses to Adalimumab up to 24%. It is of urgency to develop a cost-effective and easy-to-use ADA diagnostic kit for diagnosis of potential drug-resistance in patients treated with Adalimumab in clinic hospitals to avoid the tremendous economic and human costs to patients and health-care providers. In this study, we reported the generations of mouse monoclonal and monkey polyclonal antibodies against Adalimumab as assay standards and positive quality controls respectively. A Bridging ELISA assay was successfully developed with a limit of detection (LOD) between 22–80ng/ml. The preliminary validation of assay was carried out first with 50 normal human sera, further validated by screening the ADA in 192 serum samples from monkeys treated with or without Adalimumab. Our data showed that the Bridging ELISA kit is very sensitive, highly specific and ready for study in human clinic trials.

Effects of mindfulness-based stress reduction on anxiety, depression and immune function in patients with gastric cancer

Objective To explore the effects of mindfulness-based stress reduction (MBSR) on anxiety, depression and immune function in patients with gastric cancer. Methods The study investigated 118 patients with gastric cancer who were hospitalized from March 2011 to February 2015. The participants were randomly divided into the intervention group (n=59) and the control group (n=59). Both groups received conventional mental health education while the intervention group complemented with MBSR intervention once a week (six weeks in total). Self-rating anxiety scale (SAS) and self-rating depression scale (SDS) were used to evaluate patients′ anxiety and depression status before and after the intervention. Moreover, T-cell subsets status could reflect the immune function using phosphatase bridging ELISA. Results Before the intervention, the scores of SAS and SDS had no significant differences between the two groups(P>0.05). However, after the MBSR intervention, the scores for the intervention group significantly reduced and lower than the control group (P 0.05). Furthermore, the CD3+, CD4+, CD8+ and CD4+ /CD8+ had little differences between the two groups before the intervention(P>0.05). However, the CD3+, CD4+ and CD4+ /CD8+ for the intervention group significantly increased with CD8+ reduced after MBSR (P 0.05). Conclusions The MBSR intervention can significantly reduce the negative emotions such as anxiety and depression, and can increase the immunity in patients with gastric cancer. Key words: Anxiety; Depression; Immune function; Gastric cancer; Mindfulness-based stress reduction

Detection of adalimumab and anti-adalimumab antibodies in patients with rheumatoid arthritis: a comprehensive overview of methodology pitfalls and benefits.

About a third of patients with rheumatoid arthritis treated with adalimumab may develop anti-adalimumab antibodies. Anti-adalimumab antibodies are associated with reduced drug levels, loss of drug efficacy, clinical non-response and an increased risk of adverse effects. In case of suspected drug failure and in order to better define clinical efficacy, adalimumab as well as anti-adalimumab antibodies levels should be monitored. Sandwich or indirect enzyme-linked immunoassay is most commonly used for determining adalimumab, while bridging ELISA and antigen-binding test are most useful for determining anti-adalimumab antibodies. Most current assays cannot detect antibodies complexed with the adalimumab; however, methods for dissociation of the complexes using acid/temperature have been developed. The aim of this review is to report on the latest methodology for detecting adalimumab and anti-ADL antibodies, benefits of their detections in clinical practice, as well as expose problematic issues, such as different analytical sensitivity and specificity, standardization and validation. The main problem in measuring adalimumab or anti-ADL antibodies is high drug sensitivity, which can result in false-negative anti-ADL antibodies. Therefore, drug-tolerant assays have been developed. Cell-based assays, such as the reporter gene assay, are recommended for detection of functionally active adalimumab and their neutralizing anti-ADL antibodies.

Abstract 1411: Plasma and cerebrospinal fluid pharmacokinetics of tocilizumab in a nonhuman primate model

Abstract Background: Cytokine release syndrome (CRS) is a life-threatening complication of effective immunotherapies, particularly chimeric antigen receptor (CAR) T-cells. CRS is characterized by fever, hypotension, cardiac dysfunction, and at times end organ dysfunction and is mediated by elevated cytokines including IL-6, IFN-γ, and TNFα. A mainstay treatment for severe CRS is intravenous (IV) tocilizumab, a humanized, monoclonal antibody against membrane-associated and soluble IL-6 receptor. Most CRS symptoms resolve within hours of a single dose of IV tocilizumab in contrast to neurotoxicities, which are exacerbated or develop de novo after tocilizumab by a mechanism that is not well understood. IL-6 is elevated in the cerebrospinal fluid (CSF) of patients with CRS and neurotoxicity. The CSF penetration of tocilizumab is unknown. The objective of this study is to determine the CSF penetration of tocilizumab after IV administration in a nonhuman primate model. Methods: Four adult male rhesus monkeys (Macaca mulatta) each received 10 mg/kg tocilizumab IV (human equivalent dose of 8 mg/kg). The macaques have an implanted CNS ventricular reservoir for CSF collection and two central venous lines; one for drug administration and a second for sample collection. Serial, paired blood and CSF samples were collected 0-8 hours, 1-3 days, 5 days, & 10 days post-infusion. A fluorescence bridging ELISA was used to determine serum and CSF levels of free tocilizumab (AbD Serotec). Results: Three of the four animals were evaluable. Mean serum Cmax = 1.55 μM (1.20-2.13) and mean CSF Cmax = 0.00065 μM (0.00037-0.00083). Mean serum and CSF Tmax were 10 hours and 48 hours, respectively. Mean serum and CSF half-life were 127 and 339 hours, respectively. Serum AUC (mean) = 63 μM/hour (25-130) while CSF AUC (mean) = 0.099 μM/hour (0.07-0.125). CSF penetration was low with mean CSF:plasma AUC ratio of 0.26%. All samples and standards were plated in triplicate and had a coefficient of variation less than 20%. In one animal, tocilizumab concentrations were markedly lower with no detectable tocilizumab in the serum after 4 hours; results from this study were considered unevaluable. Conclusion: Studies in this nonhuman primate model reveal low CSF penetration of tocilizumab and delayed time to maximum concentration in the CSF, potentially explaining the persistent neurologic toxicity observed in patients with CRS treated with tocilizumab. However, tocilizumab has a long half-life in the CSF, suggesting alternate means of delivery may be beneficial. Free tocilizumab concentrations were extremely low in one out of four animals and may have been due to preexisting inhibitors. Intranasal delivery of tocilizumab may result in significantly higher CSF exposure and provides a convenient route in patients that are otherwise critically ill. Studies of intranasal tocilizumab are ongoing and will be presented. Citation Format: Anandani Nellan, Nalini Jayaprakash, Cindy McCully, Brigitte C. Widemann, Daniel W. Lee, Katherine E. Warren. Plasma and cerebrospinal fluid pharmacokinetics of tocilizumab in a nonhuman primate model. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1411.

Characterization and Application of a Unique Panel of Monoclonal Antibodies Generated against Etanercept.

The clinical response in ankylosing spondylitis (AS) patients treated with biologic agents can be influenced by pharmacokinetic variability among and within these patients. Therapeutic drug monitoring is seen as a valuable tool to improve patient care. The aim of this study was to generate a panel of mAbs toward etanercept (ETN) and to determine ETN and anti-ETN concentrations in AS patients. mAbs against ETN (MA-ETN) were generated using hybridoma technology. For quantification of ETN concentrations, a mAb-based TNF-coated ELISA and a mAb/mAb-based sandwich-type ELISA were developed. For evaluation of the anti-ETN Ab response, a bridging ELISA, as well as a functional cell-based assay, were constructed. Disease activity of the AS patients was measured with the AS Disease Activity Score (ASDAS). Active disease was defined as ASDAS ≥ 2.1. A total of 59 of 76 generated mAbs were ETN specific and were characterized further. Fifty-one mAbs revealed inhibitory properties in a cell-based assay. Analysis of serum concentrations of 21 ETN-treated AS patients with the TNF/MA-ETN68C5-HRP ELISA and the MA-ETN63C8/MA-ETN61C1-HRP ELISA revealed a good Pearson's r (+0.974) but a poor intraclass correlation coefficient (+0.528) as the result of underestimation of the values in the former ELISA. At 24 wk, ETN concentrations were similar in patients with ASDAS < 2.1 and ≥ 2.1. Anti-ETN Abs were not detected in any of the patient samples tested. In conclusion, highly sensitive mAb-based immunoassays were developed for quantification of ETN and anti-ETN concentrations. The impact of these methods needs to be evaluated further in clinical practice.

Antibodies to infliximab in Remicade-treated rheumatic patients show identical reactivity towards biosimilars

ObjectivesThe aim of this study was to determine whether antibodies to infliximab (IFX) in Remicade-treated patients cross-react with the biosimilar CT-P13.Methods250 consecutive patients with rheumatic diseases under Remicade and 77...

Immunogenicity screening assay development for a novel human-mouse chimeric anti-CD147 monoclonal antibody (Metuzumab)

The clinical effect of patient immune responses to therapeutic antibodies affect product safety and efficacy, which makes the development of valid, sensitive immune assays a key aspect of antibody drug development. In this paper, we reported the generations of mouse monoclonal and Cynomolgus monkey polyclonal antibodies against the anti-CD147 antibody (Metuzumab) as the internal standards and the positive controls. Seven mouse monoclonal antibodies were shown to recognize both (Fab)2 and full length of Metuzumab, but not the control normal human IgGs, and monoclonal anti-Metuzumab, Clone 2D9 was chosen to be used as the internal standard for anti-Metuzumab study. A Bridging ELISA assay was developed by coating the wells with the antibody drug, and the anti-drug antibody (ADA) in the animal sera were detected by enzyme-labeled antibody. Its limit of detection (LOD) was determined to be 0.39ng/ml of anti-Metuzumab antibody (ADA) with linear range between 0.39–50ng/ml and R2=0.994. For normal monkey sera, a minimal dilution was determined to be 1:80. However, very different from peptide or other protein drugs, strong interferences from the residual antibody drugs were observed from most of the testing monkey sera in the preclinical study. It was experimentally determined that the concentration of the residual antibody drug in the assay have to be lower than 1μg/ml, so the assays were carried out at 1:100 dilution of the monkey sera. In the pre-clinical study, 32 monkeys were treated with escalating doses of Metuzumab between 0, 10, 50, 200mg/kg for 13 times over 13weeks of time period. 16 of them were terminated right after the last injection, while the other 16 were rested for additional 4weeks before termination. Afraid to miss any positive response to antibody drug, sera samples were collected at six time points, including 2-, 6- and 10-weeks post 1st dose, prior to last dose, and 2-, 4-weeks into recovery. The highest positive rates were seen with the Medium- and High-dose group 2-weeks post the first injection, 6 out 8 monkeys in the High-dose were positive for free ADA. However, no significant pathologic and clinic adversary effect was observed in those monkeys.

Variability in Golimumab Exposure: A 'Real-Life' Observational Study in Active Ulcerative Colitis.

Golimumab has been approved recently to treat refractory moderate-to-severe ulcerative colitis [UC]. To date it is not clear why a considerable fraction of patients do not respond, or lose initial response, to golimumab therapy. Our aim was to investigate whether a low golimumab serum concentration and/or a positive anti-golimumab antibody status reduces the efficacy of this drug in patients with UC. Serum samples of 21 patients with moderate-to-severe UC were collected during the first 14 weeks of golimumab therapy. For measurement of golimumab serum concentrations, both a tumour necrosis factor [TNF]-coated enzyme-linked immunosorbent assay [ELISA] and a sandwich-type ELISA were developed. Anti-golimumab antibodies were measured using a bridging ELISA and a newly-developed drug-tolerant immunoassay. Clinical response and mucosal healing were assessed 14 weeks after start of treatment. Out of 21 patients, 10 [48%] reached partial clinical response at Week 14. Median [interquartile range] serum golimumab concentration was significantly higher in partial clinical responders than in non-responders: 10.0 [7.8-10.5] µg/ml versus 7.4 [4.8-8.3] µg/ml at Week 2 [p = 0.035] and 5.1 [4.0-7.9] µg/ml versus 2.1 [1.8-4.2] µg/ml at week 6 [p = 0.037]. Four out of 21 UC patients developed anti-golimumab antibodies, detectable only using a drug-tolerant immunoassay, and three had a partial clinical response at that time. Clinical non-responders had a significantly more severe colitis, indicated by a higher endoscopic Mayo score at baseline compared with partial clinical responders [p = 0.048]. Adequate exposure to golimumab drives clinical response. A worse disease at baseline influences clinical response rate negatively.

Preexisting Antibodies to an F(ab')2 Antibody Therapeutic and Novel Method for Immunogenicity Assessment.

Anti-therapeutic antibodies (ATAs) may impact drug exposure and activity and induce immune complex mediated toxicity; therefore the accurate measurement of ATA is important for the analysis of drug safety and efficacy. Preexisting ATAs to the hinge region of anti-Delta like ligand 4 (anti-DLL4) F(ab′)2, a potential antitumor therapeutic, were detected in cynomolgus monkey serum, which presented a challenge in developing assays for detecting treatment induced ATA. A total ATA assay was developed using a bridging ELISA that detected both anti-CDR and anti-framework ATA including anti-hinge reactivity. A competition assay that could detect 500 ng/mL of anti-CDR ATA in the presence of preexisting ATA was also developed to determine ATA specific to the anti-DLL4 F(ab′)2 CDR using anti-DLL4 F(ab′)2 and a control F(ab′)2. We used these assay methods in a cynomolgus monkey in vivo study to successfully evaluate total and anti-CDR ATA. The preexisting anti-hinge reactivity was also observed to a lesser extent in human serum, and a similar approach could be applied for specific immunogenicity assessment in clinical trials.

Using monoclonal antibodies as an international standard for the measurement of anti-adalimumab antibodies

Comparing studies investigating anti-drug antibody (ADA) formation is hampered by the lack of comparability between study protocols, assay formats, and standardized reference materials. In this respect, the use of an international standard would mean a major step forward. Here we compared 11 fully human monoclonal antibodies against adalimumab in two assays commonly used for ADA measurement; the bridging ELISA and the antigen binding test (ABT). Our results show non-parallel titration of the monoclonal antibodies in both assays, which we also find for polyclonal ADA sources. Moreover, we observed that the output of the bridging ELISA depends to a large degree on the affinity of the monoclonal antibody. For the ABT, results reflect a combination of affinity and avidity. This suggests that rather than reporting ADA values in nanogram per milliliter, arbitrary units may be more appropriate. Together our data highlight the difficulty of ADA standardization by identifying several pitfalls that should be taken into account when selecting a standard for ADA testing.

PTH-091 Measurement of tnf-alpha drug levels and free versus total anti-drug antibodies using three commercially available assays: Abstract PTH-091 Table 1

<h3>Introduction</h3> Commercial assays are now available for therapeutic drug monitoring (TDM) of anti-TNF drugs and antibodies (ADAb). Utility of free versus total ADAb assays remains debatable, further complicated by lack of assay standardisation. Here we report analytical comparison of 3 commercially available assays for Infliximab (IFX) and Adalimumab (ADAL) drug levels (DL) and ADAb. <h3>Method</h3> Prospective evaluation of IFX and ADAL DL and ADAb was performed using our local LISA-TRACKER (LT) assay automated on e-Robot in IBD patients. Samples were also analysed by Immundiagnostik (IM, Germany) and Promonitor (PM, Spain) ELISA automated on Grifols Triturus. LT and PM utilises a specific bridging ELISA to quantitatively measure free-ADAb whereas IM utilises a dissociation step to enable detection of total-ADAb generating semi-quantitative results. IFX assays measure free drug but differ in microtitre plate coating and secondary detection reagents. Data was analysed using Passing Bablok and bias plots. LT and PM kits were provided at no cost. <h3>Results</h3> Summary of DL comparisons shown below: Samples analysed in different batches showed different kit biases against each other for IFX. Batch 1 showed that LT assay had 43.9% positive bias against ID kit whereas batch 2 demonstrated –26% negative bias. Both LT and ID kits used had different lot numbers. This change in bias was not observed in ADAL assays which showed consistent and systematic bias. PM kit showed concentration dependent bias changes within the same assay. 4 patients tested (n = 79) IFX ADAb positive with undectable DL with one exception where total/free ADAb was negative using ID and PM assays. A further 17 patients tested total ADAb positive using IM with detectable DL (0.5–9.2 ug/mL). 1 patient tested ADAL ADAb positive using LT, PM and ID assays however ID and PM assays produced positive resuts on a further 4 specimens, all with ADAL DL &gt;5 ug/mL. <h3>Conclusion</h3> Although commercial assays are now available, our data highlights the need for assay standardisation. Free ADAb assays were in agreement however ADAb positivity was higher using ID assay. Significance of total ADAb positivity is unknown. Results for both IFX and ADAL confirm that DL are not transferable from one assay to another and as such, common therapeutic cut-offs will not be applicable. Further work is warranted to establish the cause of batch-to-batch variation observed. <h3>Disclosure of interest</h3> Z. Arkir: None Declared, N. Unsworth: None Declared, B. Warner: None Declared, G. Richards: None Declared, P. Irving Speaker Bureau of: MSD, Abbvie and Takeda.

Prevalence of TNF-α blocker immunogenicity in psoriatic arthritis.

The longterm use of tumor necrosis factor (TNF)-α blockers is limited by the formation of neutralizing antibodies. To the best of our knowledge, immunogenicity in psoriatic arthritis (PsA) has not been investigated in depth. Our objective was to evaluate the prevalence and significance of TNF-α blocker immunogenicity in PsA. Consecutive patients with PsA treated with either infliximab (IFX), adalimumab (ADA), or etanercept (ETN) > 3 months participated in our cross-sectional study. Their demographic and clinical characteristics, skin and joint disease activity, and records of use of methotrexate (MTX) and other medications were collected. Drug levels (ELISA) and antidrug antibodies (ADAb; Bridging ELISA) were evaluated before the next injection or infusion. A total of 93 patients with PsA were recruited (48 receiving ADA, 24 IFX, and 21 ETN), with a mean age of 53 years (range 21-83 yrs), composed of 53% women. One-fourth of the patients were concomitantly treated with MTX. Altogether, 77% of the patients demonstrated therapeutic drug levels. High levels of ADAb were found in 29% of patients taking ADA, 21% taking IFX, and 0% taking ETN. ADAb significantly correlated with lower drug levels, higher 28-joint Disease Activity Scores, and higher global assessments. MTX use correlated significantly with a lower prevalence of ADAb. Significant levels of ADAb were present in up to 29% of patients with PsA treated with ADA or IFX. ADAb clearly correlated with low therapeutic drug levels and higher disease activity variables. The use of MTX significantly decreased ADAb prevalence, and its use should be strongly considered in combination with TNF-α blocker antibodies in patients with PsA.

Challenges in keeping pace with immunogenicity advances throughout the clinical development of a biological therapeutic.

Progress in the assessment of immunogenicity over the past decade has been rapid; therapeutics in late-phase clinical studies today are subject to antidrug antibody analyses that have evolved considerably since the start of their clinical development. Data from bridging and direct ELISA methods used for the analysis of antidrug antibodies generated in response to an antibody therapeutic will be presented. The case study will highlight some of the challenges faced and the learning points gained from supporting immunogenicity analysis for these therapeutics.

SP0127 Basic Concepts: Analysis of Anti-Drug Antibodies and Serum Concentrations of Biologicals

BackgroundTreatment with biological drugs in the last fifteen years has changed the therapy of most inflammatory diseases, mainly in the field of rheumatology. The use of these drugs has supposed...

OP0253 The Early Infliximab Levels Monitoring Can Predict the Developement of Anti-Drug Antibodies in a Cohort of Rheumatoid Arthritis Patients Treated with Infliximab

BackgroundThere is strong evidence that correlates the antibodies to infliximab appearance (ATI) with a poor clinical response, but for now sparse literature has demonstrated the predictive value of early serum...

Sa1222 Fecal Lactoferrin, Calprotectin, PMN-Elastase, CRP and White Blood Count As Indicator for Clinical Course of Disease and Mucosal Healing in Patients With Ulcerative Colitis: A Prospective 12-Month Monitoring Study

and AAA levels (measured with a previously described ELISA and bridging ELISA respectively) were available from routine follow-up at our IBD unit. Twenty patients (17 CD and 3 UC) were identified of which 89 serum samples were retrospectively tested in a functional TNF-responsive cell based assay (CBA) to determine neutralising capacity of the AAA. MA-ADM6A10, a monoclonal antibody with a high affinity and specificity for adalimumab was used as standard in CBA, hence AAA values were expressed towards the response of this MA. Neutralising AAA levels <0.132mg/ml were regarded as undetectable. Results: Testing with CBA revealed neutralising AAA in 48/89 samples (53.9%) thereby classifying all 20 patients as positive for neutralising AAA on at least one time point during follow up. CBA detected AAA in 15 samples that were negative in bridging ELISA, thereby reclassifying 3 patients as AAA positive. Median (inter quartile range) levels of AAA in ELISA were 0.20 mg/ml equivalents (0.20 0.39) and 0.21 mg/ml (0.13 1.4) in CBA. Of the patients developing neutralising AAA 15/20 patients (75%) discontinued adalimumab treatment or were still treated with adalimumab but were not in remission at the end of follow-up. Conclusions: Functional testing with a CBA revealed neutralising AAA in 5% of IBD patients treated with maintenance adalimumab. CBAwas more sensitive than ELISA but both assays cannot detect AAA in presence of drug and therefore the amount of AAA positive patients is underestimated. However, this study demonstrates that the development of detectable neutralising AAA although not frequent is detrimental for the patient’s clinical outcome.

Significant associations of antidrug antibody levels with serum drug trough levels and therapeutic response of adalimumab and etanercept treatment in rheumatoid arthritis

ObjectiveTo evaluate the associations between (1) antidrug antibody (ADAb) and therapeutic response, (2) ADAb and serum drug trough levels and (3) serum drug levels and therapeutic responses in rheumatoid arthritis...

A Simple and Fast Non-Radioactive Bridging Immunoassay for Insulin Autoantibodies

Type 1 diabetes (T1D) is an autoimmune disease which results from the destruction of pancreatic beta cells. Autoantibodies directed against islet antigens are valuable diagnostic tools. Insulin autoantibodies (IAAs) are usually the first to appear and also the most difficult to detect amongst the four major islet autoantibodies. A non-radioactive IAA bridging ELISA was developed to this end. In this assay, one site of the IAAs from serum samples is bound to a hapten-labeled insulin (GC300-insulin), which is subsequently captured on anti-GC300 antibody-coated 96-well plates. The other site of the IAAs is bound to biotinylated insulin, allowing the complex to be detected by an enzyme-streptavidin conjugate. In the present study, 50 serum samples from patients with newly diagnosed T1D and 100 control sera from non-diabetic individuals were analyzed with our new assay and the results were correlated with an IAA radioimmunoassay (RIA). Using IAA bridging ELISA, IAAs were detected in 32 out of 50 T1D children, whereas with IAA RIA, 41 out of 50 children with newly diagnosed T1D were scored as positive. In conclusion, the IAA bridging ELISA could serve as an attractive approach for rapid and automated detection of IAAs in T1D patients for diagnostic purposes.

PWE-109 Measurement of Infliximab and Anti-Infliximab Antibodies - Analytical Aspects and Clinical Implications

IntroductionCommercial assays for monitoring Infliximab (IFX) and anti-drug antibodies (ADAb) are currently available. The heterogeneity of these assays and lack of standardisation of ADAb may cause difficulties in interpretation, in...