Detection of anti-infliximab antibodies is impacted by antibody titer, infliximab level and IgG4 antibodies: a systematic comparison of three different assays.
BACKGROUND
There is scant information on the accuracy of different assays used to measure anti-infliximab antibodies (ADAs), especially in the presence of detectable infliximab (IFX). We thus aimed to evaluate and compare three different assays for the detection of IFX and ADAs and to clarify the impact of the presence of circulating IFX on the accuracy of the ADA assays.


METHODS
Blood samples from 79 ulcerative colitis (UC) patients treated with infliximab were assessed for IFX levels and ADAs using three different assays: an in-house assay and two commercial kits, Immundiagnostik and Theradiag. Sera samples with ADAs and undetectable levels of IFX were spiked with exogenous IFX and analyzed for ADAs.


RESULTS
The three assays showed 81-96% agreement for the measured IFX level. However, the in-house assay and Immundiagnostik assays detected ADAs in 34 out of 79 samples, whereas Theradiag only detected ADAs in 24 samples. Samples negative for ADAs with Theradiag, but ADA-positive in both the in-house and Immundiagnostik assays, were positive for IFX or IgG4 ADAs. In spiking experiments, a low concentration of exogenous IFX (5 µg/ml) hampered ADA detection with Theradiag in sera samples with ADA levels of between 3 and 10 µg/ml. In the Immundiagnostik assay detection interference was only observed at concentrations of exogenous IFX higher than 30 µg/ml. However, in samples with high levels of ADAs (>25 µg/ml) interference was only observed at IFX concentrations higher than 100 µg/ml in all three assays. Binary (IFX/ADA) stratification of the results showed that IFX+/ADA- and IFX-/ADAs+ were less influenced by the assay results than the double-positive (IFX+/ADAs+) and double-negative (IFX-/ADAs-) combination.


CONCLUSIONS
All three methodologies are equally suitable for measuring IFX levels. However, erroneous therapeutic decisions may occur when patients show double-negative (IFX-/ADAs-) or double-positive (IFX+/ADAs+) status, since agreement between assays is significantly lower in these circumstances.
Department of Pharmacology and Therapeutics, Faculty of Medicine, University of Porto, Porto, Portugal MedInUP, Centre for Drug Discovery and Innovative Medicines, University of Porto, 4200 Porto, Portugal.
Joana
J
Afonso
Gastroenterology Department, Centro Hospitalar São João, Porto, Portugal.
Susana
S
Lopes
Gastroenterology Department, Hospital de Braga, Braga, Portugal.
Raquel
R
Gonçalves
Gastroenterology Department, Centro Hospitalar do Algarve, Faro, Portugal.
Paulo
P
Caldeira
Gastroenterology Department, Centro Hospitalar do Porto, Porto, Portugal.
Paula
P
Lago
Gastroenterology Department, Centro Hospitalar do Algarve, Portimão, Portugal Departament of Medicine e Medical Biosciences, University of Algarve, Faro, Portugal.
Helena
H
Tavares de Sousa
Gastroenterology Department, Centro Hospitalar de Lisboa, Lisboa, Portugal.
Jaime
J
Ramos
Gastroenterology Department, Centro Hospitalar Lisboa Norte, Lisboa, Portugal.
Ana Rita
AR
Gonçalves
Gastroenterology Department, Hospital de S. Teotónio, Viseu, Portugal.
Paula
P
Ministro
Gastroenterology Department, Instituto Português de Oncologia de Lisboa, Lisboa, Portugal.
Isadora
I
Rosa
Gastroenterology Department, Hospital Garcia de Orta, Almada, Portugal.
Ana Isabel
AI
Vieira
Gastroenterology Department, Centro Hospitalar São João, Porto, Portugal.
Rosa
R
Coelho
Gastroenterology Department, Centro Hospitalar São João, Porto, Portugal.
Patrícia
P
Tavares
Gastroenterology Department, Hospital de Braga, Braga, Portugal.
João
J
Soares
Gastroenterology Department, Centro Hospitalar do Algarve, Faro, Portugal.
Ana Lúcia
AL
Sousa
Gastroenterology Department, Centro Hospitalar de Lisboa, Lisboa, Portugal.
Diana
D
Carvalho
Gastroenterology Department, Hospital de S. Teotónio, Viseu, Portugal.
Paula
P
Sousa
Gastroenterology Department, Instituto Português de Oncologia de Lisboa, Lisboa, Portugal.
João Pereira
JP
da Silva
Gastroenterology Department, Hospital Garcia de Orta, Almada, Portugal.
Tânia
T
Meira
Department of Pharmacology and Therapeutics, Faculty of Medicine, University of Porto, Porto, Portugal MedInUP, Centre for Drug Discovery and Innovative Medicines, University of Porto, 4200 Porto, Portugal.
Filipa
F
Silva Ferreira
Department of Pharmacology and Therapeutics, Faculty of Medicine, University of Porto, Porto, Portugal.
Cláudia Camila
CC
Dias
Gastroenterology Department, Rambam Health Care Campus and Bruce Rappaport School of Medicine, Technion Israel Institute of Technology, Israel.
Yehuda
Y
Chowers
IBD Service, Department of Gastroenterology, Sheba Medical Center and Sackler School of Medicine, Tel-Aviv University, Israel.
Shomron
S
Ben-Horin
Gastroenterology Department, Medical School, Centro Hospitalar São João, Alameda Prof. Hernâni Monteiro, 4200-319 Porto, Portugal.
Fernando
F
Magro
anti-Infliximab antibody methodologies; anti-infliximab antibodies; infliximab trough levels; therapeutic drug monitoring