Fluorescence-Based Lateral Flow Immunoassay for Quantification of Infliximab: Analytical and Clinical Performance Evaluation

Abstract

Background: Therapeutic drug monitoring of infliximab (IFX) can improve treatment outcomes; however, the temporal gap between drug concentration monitoring and subsequent availability restricts its practical application. To address this issue, an automated monitoring method, AFIAS IFX, was developed to rapidly and accurately analyze IFX concentration in blood. The analytical and clinical performances of this method were assessed to establish its clinical utility.

Methods: The analytical performance of AFIAS IFX was evaluated according to Clinical and Laboratory Standard Institute guidelines. For clinical validation, AFIAS IFX was compared with 3 established enzyme-linked immunosorbent assay kits (LISA TRACKER, RIDASCREEN, and ImmunoGuide) using 100 consecutive samples from 28 patients treated with IFX. Passing-Bablok regression and Bland-Altman analyses were performed to compare the methods.

Results: The detection and quantification limits of AFIAS IFX were 0.12 and 0.20 mcg/mL, respectively. Furthermore, AFIAS IFX analyzed samples within 10 minutes for concentrations up to 50 mcg/mL, exhibiting reproducibility (coefficient of variation [CV] ≤7.8%) and accuracy (recovery 98%-101%) with serum, plasma, and whole blood samples. Clinically, it exhibited a good correlation with the 3 established enzyme-linked immunosorbent assay kits. For patients treated with Remicade (IFX), the Passing-Bablok regression slope was 1.001-1.259, with a mean difference of -1.48 to 0.28 mcg/mL. For patients treated with CT-P13, the Passing-Bablok regression slope was 0.974-1.254, with a mean difference of -2.44 to 0.15 mcg/mL.

Conclusions: AFIAS IFX, a novel fluorescence-based lateral flow assay, exhibited excellent performance in analyzing IFX trough levels and is a potentially powerful tool for therapeutic drug monitoring in clinical settings, with opportunities for further development.

FIGURE 1.

Diagram illustrating the mechanism of a fluorescence-based lateral flow immunoassay. A, Formation of sandwich immunocomplexes between IFX and ADA. B, Immobilization and fluorescence signal detection of sandwich immunocomplexes.

FIGURE 2.

Passing–Bablok regression analysis of quantifiable (n = 50) IFX drug levels analyzed using 3 different ELISA kits, (A) LISATracker, (B) RIDAScreen, and (C) ImmunoGuide, compared with AFIAS IFX. The data show the results of samples (n = 50) from treated patients treated with Remicade. The regression equation (blue solid line) reveals a constant difference (the regression line intercept and slope). The red dotted lines indicate the 95% CI. The green dotted line is the identity line.

FIGURE 3.

Passing–Bablok regression analysis of quantifiable (n = 50) IFX drug levels analyzed using 3 different ELISA kits, (A) LISATracker, (B) RIDAScreen, and (C) ImmunoGuide, compared with AFIAS IFX. The data show the results of samples (n = 50) from patients treated with CT-P13. The regression equation (blue solid line) reveals a constant difference (the regression intercept and slope). The red dotted lines indicate the 95% CI. The green dotted line is the identity line.