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Related Experiment Videos

Engineered recombinant single-chain fragment variable antibody for immunosensors.

Zhihong Shen1, Raymond L Mernaugh, Heping Yan

  • 1Department of Chemistry, Oakland University, Rochester, Michigan 48309, USA.

Analytical Chemistry
|November 1, 2005
PubMed
Summary

Engineered antibodies with histidines in the linker peptide demonstrate effective antigen detection. This novel piezoimmunosensor maintains antibody function and offers sensitive detection in complex samples.

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Area of Science:

  • Biotechnology
  • Immunosensor Technology
  • Protein Engineering

Background:

  • Recombinant single-chain fragment variable (scFv) antibodies are crucial for developing sensitive diagnostic tools.
  • Optimizing the orientation and functionality of scFv antibodies on sensor surfaces is key for high-performance immunosensors.

Purpose of the Study:

  • To engineer a novel scFv antibody with histidines in the linker peptide for enhanced piezoimmunosensor development.
  • To characterize the performance of the histidine-containing scFv in a quartz crystal microbalance (QCM) sensor for antigen detection.

Main Methods:

  • Engineering of a recombinant scFv antibody (A10B) with two histidines within the linker peptide.
  • Development and characterization of a gold piezoimmunosensor utilizing the engineered A10B scFv.

Related Experiment Videos

  • Quantification of antigen detection sensitivity and binding kinetics using the scFv-modified QCM sensor.

    • Main Results:

    • The A10B scFv demonstrated correct orientation, retained antigen-binding activity, and high surface coupling on the piezoimmunosensor.
    • The histidine incorporation in the linker was well-tolerated and did not impede scFv antigen-binding activity.
    • The sensor achieved a sensitivity of 2.3 +/- 0.15 nM for antigen detection in crude serum, with a linear range from 2.3 x 10(-9) to 3.3 x 10(-8) M.

    Conclusions:

    • Histidine incorporation into the scFv linker is a viable strategy for developing functional piezoimmunosensors.
    • The developed piezoimmunosensor offers specific and sensitive antigen detection, suitable for complex biological samples.
    • The sensor platform enables detailed study of antigen-antibody binding kinetics and thermodynamics.