Development and validation of an optical biosensor for rapid monitoring of adalimumab in serum of patients with Crohn's disease.

Therapeutic drug monitoring of adalimumab is recommended to improve therapeutic outcome in patients with Crohn's disease. Performing an ELISA requires a rather long time-to-result and the necessity of collecting multiple samples to decrease the cost per adalimumab determination. In this study, we aim to develop and validate a rapid assay suitable for measuring a single adalimumab serum sample using a fiber-optic surface plasmon resonance (FO-SPR) based sensor. Therefore, we have immobilized MA-ADM28B8 as capture antibody on an FO-probe and conjugated MA-ADM40D8 as detecting antibody to gold nanoparticles. A dose-response curve ranging from 2.5 to 40 ng/mL adalimumab was obtained in 1/400 diluted serum. Serum samples of patients with adalimumab concentrations between 1 and 16 µg/mL were measured whereas the negative control, a sample spiked with infliximab at a concentration of 16 µg/mL, showed no significant signal. Using a pre-functionalized FO-probe, the technology requires less than 45 minutes for measuring a single sample. Comparison of measurements between the biosensor and the ELISA revealed an excellent agreement with a Pearson r coefficient of 0.99 and an intra-class coefficient of 0.99. The reduced assay time and the possibility of measuring a single sample are major advantages compared to the ELISA. The developed and validated optical adalimumab biosensor could be a valuable point-of-care diagnostic tool for adalimumab quantification in patients with Crohn's disease.

Immunoassay for Detection of Infliximab in Whole Blood Using a Fiber-Optic Surface Plasmon Resonance Biosensor.

Monitoring the concentration of a therapeutic drug antibody, infliximab (IFX), is recommended for enhancing its efficacy in patients with inflammatory bowel disease (IBD). However, IFX concentrations are currently determined in patients' serum/plasma, which requires sample preparation from blood, hence hampering the turnaround time. In this paper, we present a short immunoassay (10 min) using a fiber-optic surface plasmon resonance (FO-SPR) biosensor for detection of IFX spiked in 100-fold diluted serum, plasma, and whole blood. The calculated limits of detection (LOD) based on calibration curves were 1.42, 1.00, and 1.34 ng/mL, respectively, which coincides with expected IFX concentrations in diluted samples from IBD patients. A linear correlation was established among different matrixes, indicating that the matrix effect was insignificant. The established point-of-care (POC) FO-SPR bioassay was also used to measure IFX in 100-fold diluted extracts of dried blood spots (DBS), and LOD achieved was below 2 ng/mL. Although DBS might be ideal for POC, this is the first report of using an SPR biosensor for measuring DBS samples. Finally, the POC FO-SPR immunoassay was validated by using matching serum and plasma samples from five IBD patients. A Pearson correlation of 0.968 was obtained between serum and plasma samples. IFX concentrations determined with FO-SPR were compared to a clinically validated enzyme-linked immunosorbent assay (ELISA), resulting in excellent Pearson correlation and intraclass correlation coefficient, both being 0.99 for serum and plasma samples. In conclusion, this paper demonstrates that our FO-SPR biosensor can be used as a true POC diagnostic tool for determining IFX concentrations in a variety of matrixes.

Optical Manipulation of Single Magnetic Beads in a Microwell Array on a Digital Microfluidic Chip.

The detection of single molecules in magnetic microbead microwell array formats revolutionized the development of digital bioassays. However, retrieval of individual magnetic beads from these arrays has not been realized until now despite having great potential for studying captured targets at the individual level. In this paper, optical tweezers were implemented on a digital microfluidic platform for accurate manipulation of single magnetic beads seeded in a microwell array. Successful optical trapping of magnetic beads was found to be dependent on Brownian motion of the beads, suggesting a 99% chance of trapping a vibrating bead. A tailor-made experimental design was used to screen the effect of bead type, ionic buffer strength, surfactant type, and concentration on the Brownian activity of beads in microwells. With the optimal conditions, the manipulation of magnetic beads was demonstrated by their trapping, retrieving, transporting, and repositioning to a desired microwell on the array. The presented platform combines the strengths of digital microfluidics, digital bioassays, and optical tweezers, resulting in a powerful dynamic microwell array system for single molecule and single cell studies.

Fiber optic-SPR platform for fast and sensitive infliximab detection in serum of inflammatory bowel disease patients.

Infliximab (IFX) is a therapeutic monoclonal antibody used for treating patients with inflammatory bowel disease (IBD). In order to improve therapeutic outcomes it is recommended to monitor IFX trough concentrations. Although ELISA is currently widely used for this purpose, this method is not suitable for single patient testing. In this paper we describe the development of a fast bioassay for determining IFX concentration in serum using an in-house developed fiber-optic surface plasmon resonance (FO-SPR) biosensor. Studies were first conducted to optimize covalent immobilization of the IFX-specific antibody on the sensor surface as well as to select an optimal blocking buffer for restraining the non-specific binding. In order to reach clinically relevant sensitivity for detecting IFX in patients' serum, the SPR signal was amplified by employing gold nanoparticles functionalized with another set of IFX specific antibodies. Using the optimized sandwich bioassay, calibration curves were made with series of IFX concentrations spiked in buffer and 100-fold diluted serum, reaching the limit of detection of 0.3 and 2.2ng/ml, respectively. The established bioassay was finally validated using five IFX treated IBD patients samples. Results from the FO-SPR platform were compared with an in-house developed, clinically validated ELISA resulting in excellent Pearson and intraclass correlation coefficient of 0.998 and 0.983, respectively. Furthermore, the assay time of the FO-SPR platform was significantly reduced compared to ELISA, demonstrating the potential of this platform to be used as a point-of-care diagnostic tool for improving therapeutic outcomes of IBD patients.