Label-free biochemical detection coupled on-line to liquid chromatography

Area of Science:

• Analytical Chemistry
• Biotechnology
• Environmental Science

Background:

• Pesticide detection requires sensitive and specific analytical methods.
• High-Performance Liquid Chromatography (HPLC) offers separation power, while biosensors provide specificity.
• Label-free optical biosensors offer advantages in real-time monitoring without the need for labels.

Purpose of the Study:

• To develop and validate an on-line coupled system integrating HPLC with a label-free optical biosensor.
• To assess the system's capability for sensitive pesticide detection using an antibody-based approach.
• To characterize the binding affinity of antibodies to pesticides.

Main Methods:

• Coupling of a label-free optical biosensor with an HPLC system.
• Utilizing a highly cross-reactive antibody against the pesticide isoproturon as a model affinity protein.
• Quantifying antigen presence by measuring the reduction in antibody binding to the transducer using a Savitzky-Golay algorithm.

Main Results:

• The binding strength of the antibody to pesticides was successfully characterized.
• The on-line system detected the presence of antigens by a quantifiable reduction in antibody binding.
• Achieved limits of detection in the femtomole range without preconcentration.

Conclusions:

• The on-line coupling of HPLC and a label-free optical biosensor is a powerful approach for sensitive pesticide analysis.
• The developed method demonstrates comparable sensitivity to fluorescence-based detection methods.
• This integrated system offers a promising tool for real-time, specific, and sensitive detection of pesticides.