Filtern
Dokumenttyp
- Zeitschriftenartikel (1)
- Vortrag (1)
Sprache
- Englisch (2)
Schlagworte
- Automation (1)
- Bead injections (1)
- Biomarkers (1)
- Carbamazepine (1)
- ELISA (1)
- Immunoassay (1)
- Lab-on-valve (1)
- Mesofluidics (1)
Organisationseinheit der BAM
Eingeladener Vortrag
- nein (1)
The lab-on-valve (LOV) is a mesofluidic platform that has been recently exploited for
the automation and miniaturization of bioanalytical assays, resorting namely to
molecular recognition schemes based on immunosensing. Due to its high versatility
for reagent accommodation, it is possible to establish immunoassays under several
formats (eg. direct competitive ELISA, sandwich ELISA or even label-free immunoaffinity
chromatography). For instance, the LOV has been used as a manifold for
UV-vis micro-Bead Injection Spectroscopy (μ-BIS), a technique that involves the
quantification of the target analyte by direct measurement on the surface of a solid
phase capable of retaining the target analyte by molecular recognition.
The μ-BIS-LOV strategy affords several analytical advantages, namely short time-toresult
intervals (3 to 15 min), low sample volume (1-20 μL), automated solution handling
and washing steps, downscaling of reagents’ consumption, low-cost analysis
and little generation of waste. Additionally, the solid support is renewed before each
determination, minimizing surface fouling, cross-contamination issues and functional
group deactivation. No sample clean-up steps are required because interferences
are separated from the target analyte upon quantification mediated by a molecular
recognition element attached to the micro-bead column. The portability of the LOV
device makes it compatible with point-of-care testing.
To our knowledge, this technique has been mainly employed for the evaluation and
optimization of bioaffinity processes, but its potential for clinical and environmental
analysis remains underexploited. Hence, in this communication, different immunosensing
strategies using the LOV platform will be addressed, namely the determination
of autoimmune IgG in human serum, and the assessment of drug (carbamazepine)
levels in wastewater samples.
Immunochemical techniques are the workhorse for sample enrichment and detection of a large variety of analytes. In contrast to classical microtiter plate-based assays, microparticles are a next generation solid support, as they promote automation of immunoassays using flow-based techniques. Antibody immobilization is a crucial step, as these reagentsmare expensive, and inefficient coupling can result in low sensitivities. This paper proposes a general procedure for efficient immobilization of antibodies onto TentaGel particles, via Nhydroxysuccinimide chemistry. The goal was the preparation of solid supports with optimum immunorecognition, while increasing the sustainability of the process. The influence of buffer composition, activation and coupling time, as well as the amount of antibody on the immobilization efficiency was investigated, resorting to fluorophore-labeled proteins and fluorescence imaging. Buffer pH and activation time are the most important parameters for efficient coupling. It is demonstrated, that the hydrolysis of N-hydroxysuccinimide esters occurs at similar rates as in solution, limiting the utilizable time for coupling. Finally, applicability of the generated material for automated affinity extraction is demonstrated on the mesofluidic platform lab-on-valve.