Abstract
A stochastic reaction–diffusion model was developed to describe the binding of labeled monoclonal antibodies (mAbs) to CD4 receptors on the surface of T cells. The mAbs diffused to, adsorbed on, and underwent monovalent and bivalent binding to CD4 receptors on the cell surface. The model predicted the time-dependent nature of all populations involved in the labeling process. At large time, the populations reached equilibrium values, giving the number of antibodies bound to the T cell (ABC) defined as the sum of monovalently and bivalently bound mAbs. The predicted coefficient of variation (CV%) of the (ABC) values translated directly to a corresponding CV% of the measured mean fluorescence intensity (MFI). The predicted CV% was about 0.2% from the intrinsic fluctuations of the stochastic reaction process, about 5% after inclusion of the known fluctuations in the number of available CD4 receptors, and about 11% when fluctuations in bivalent binding affinity were included. The fluorescence detection process is expected to contribute approximately 7%. The abovementioned contributions to CV% sum up to approximately 13%. Work is underway to reconcile the predicted values and the measured values of 17% to 22%.
Highlights
Flow cytometers measure mean fluorescence intensity (MFI) emitted from cells with antibodies that contain conjugated fluorophores
The results of the stochastic simulation were compared to the measurements of coefficient of variation (CV)% of MFI shown in Figure 1, which gives CV% as a function of label load on the T cells, as given by MFI
A stochastic reaction–diffusion model was developed to describe the binding of labeled monoclonal antibodies to CD4 receptors on the surface of T cells
Summary
Flow cytometers measure mean fluorescence intensity (MFI) emitted from cells with antibodies that contain conjugated fluorophores. The antibodies bind to specific receptors on the surface of the cell, and, with proper calibration, the MFI signal can yield the number of antibodies bound to the cell (ABC) [1]. Another property of the observed fluorescence signals is the coefficient of variation (CV), defined as the standard deviation (SD) of the fluorescence signals divided by the MFI. Of special concern is the variability of the process of labeling T cells with monoclonal antibodies (mAbs) for the CD4 receptor This manuscript describes a model for the labeling process of T cells. This is less than the observed CV% of 17% to 22%
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