Affinity measurement of single chain antibodies: a mathematical method facilitated by statistical software SigmaPlot

Area of Science:

• Biochemistry
• Immunology
• Biotechnology

Background:

• Single chain antibodies (scFv) exhibit distinct antigen-binding properties compared to full-length antibodies due to their monovalent nature.
• Understanding the affinity of scFv is critical for their application in diagnostics and therapeutics.

Purpose of the Study:

• To introduce a novel mathematical formula for estimating the affinity of single chain antibodies.
• To provide a practical and accessible method for scFv affinity determination in standard biological laboratories.

Main Methods:

• Utilized the law of mass action to model antibody-antigen interactions.
• Employed optical density (OD) measurements and sigmoid curve analysis to determine antibody concentrations at OD-50% points.
• Validated the calculated affinity against surface plasmon resonance (SPR) data.

Main Results:

• A formula for estimating single chain antibody affinity was derived based on OD-50% points.
• The method demonstrated accuracy comparable to surface plasmon resonance.
• A software-adaptable format of the equation was presented to simplify calculations.

Conclusions:

• The developed formula offers a precise and cost-effective method for determining single chain antibody affinity.
• This technique eliminates the need for specialized equipment, making scFv affinity assessment widely accessible.
• The findings facilitate the development and optimization of scFv-based biotechnological tools.