Flow Cytometry-Based Rapid Assay for Antigen Specific Antibody Relative Affinity in SRBC-Immunized Mouse Models

Sheep red blood cells (SRBC) has a long history as a classical T-cell dependent (TD) antigen. Due to its cost-effectiveness, easy accessibility, and ability to elicit a robust antibody immune response, SRBC continues to be widely used in studies related with humoral immunity modulation, vaccine development, and immunoactivity/immunotoxicity testing of bioactive agents. However, detecting the relative affinity levels of SRBC-specific antibodies in SRBC-immunized animal models remains challenging. Using flow cytometry, we established a detection system capable of quickly and accurately assessing the SRBC-specific antibody relative affinity levels in humoral samples (e.g., serum, tissue fluid) of SRBC-immunized mouse models. We further validated this method using affinity maturation-deficient mice, demonstrating that this method can distinguish affinity levels of the antibodies from different samples. This approach is simple and efficient, providing an accurate and effective technological solution for research on mechanisms of humoral immunity, antibody affinity maturation, vaccine response, and immunoactivity/immunotoxicity testing.