Estimation of antigenic tetanus toxoid extracted from biodegradable microspheres

Area of Science:

• Biomaterials Science
• Vaccine Development
• Analytical Chemistry

Background:

• Poly (lactic/glycolic acid) microspheres are explored as a novel delivery system for single-dose tetanus toxoid (TT) vaccines.
• Accurate quantification of TT loading within microspheres is crucial for vaccine efficacy and immunogenicity.
• Existing assay methods lack the sensitivity required for low TT concentrations in microspheres.

Purpose of the Study:

• To develop a highly sensitive assay for quantifying tetanus toxoid (TT) loading in poly (lactic/glycolic acid) microspheres.
• To establish an effective antigen extraction procedure that preserves TT antigenicity.
• To correlate TT loading with in vivo immunogenic responses.

Main Methods:

• Development of a monoclonal antibody (MAb) capture ELISA (MAp capture ELISA) for sensitive TT detection.
• Optimization of an antigen extraction protocol to maximize TT recovery without compromising antigenicity.
• Quantification of TT loading in various microsphere preparations using the developed assay and protein micro-BCA assay.

Main Results:

• The MAp capture ELISA demonstrated high sensitivity (0.001 Lf/ml) and was approximately 250-fold more sensitive than single-site ELISA.
• The extraction procedure yielded high antigen recovery (90.6 +/- 3.39%) and preserved TT antigenicity.
• TT content in microsphere preparations varied significantly (up to 400-fold), with higher loading correlating to greater peak immune response.

Conclusions:

• The developed MAp capture ELISA is a valuable tool for accurately determining TT loading in microspheres.
• TT loading is a critical factor influencing the peak immunogenic response.
• Further studies combining this assay with release rate and excipient data may predict in vivo vaccine performance.