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Probabilistic Modeling of Rosette Formation

Mian Long ^*, Juan Chen ^*, Ning Jiang , Periasamy Selvaraj , Rodger P. McEver  ^¶ and Cheng Zhu 

^* National Microgravity Laboratory, Institute of Mechanics, Chinese Academy of Sciences, Beijing 100080, People's Republic of China; Coulter Department of Biomedical Engineering, Georgia Institute of Technology, Atlanta, Georgia 30332; Department of Pathology and Laboratory Medicine, Emory University School of Medicine, Atlanta, Georgia 30322; and Cardiovascular Biology Research Program, Oklahoma Medical Research Foundation, and ^¶ Department of Biochemistry and Molecular Biology and Oklahoma Center for Medical Glycobiology, University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma 73104

Correspondence: Address reprint requests to: Dr. Mian Long, National Microgravity Laboratory, Institute of Mechanics, Chinese Academy of Sciences, Beijing 100080, People's Republic of China. Tel.: 86-10-6261-3540; Fax: 86-10-6261-3540; E-mail: mlong{at}imech.ac.cn.

Rosetting, or forming a cell aggregate between a single target nucleated cell and a number of red blood cells (RBCs), is a simple assay for cell adhesion mediated by specific receptor-ligand interaction. For example, rosette formation between sheep RBC and human lymphocytes has been used to differentiate T cells from B cells. Rosetting assay is commonly used to determine the interaction of Fc -receptors (FcR) expressed on inflammatory cells and IgG coated on RBCs. Despite its wide use in measuring cell adhesion, the biophysical parameters of rosette formation have not been well characterized. Here we developed a probabilistic model to describe the distribution of rosette sizes, which is Poissonian. The average rosette size is predicted to be proportional to the apparent two-dimensional binding affinity of the interacting receptor-ligand pair and their site densities. The model has been supported by experiments of rosettes mediated by four molecular interactions: FcRIII interacting with IgG, T cell receptor and coreceptor CD8 interacting with antigen peptide presented by major histocompatibility molecule, P-selectin interacting with P-selectin glycoprotein ligand 1 (PSGL-1), and L-selectin interacting with PSGL-1. The latter two are structurally similar and are different from the former two. Fitting the model to data enabled us to evaluate the apparent effective two-dimensional binding affinity of the interacting molecular pairs: 7.19 x 10^–5 µm⁴ for FcRIII-IgG interaction, 4.66 x 10^–3 µm⁴ for P-selectin-PSGL-1 interaction, and 0.94 x 10^–3 µm⁴ for L-selectin-PSGL-1 interaction. These results elucidate the biophysical mechanism of rosette formation and enable it to become a semiquantitative assay that relates the rosette size to the effective affinity for receptor-ligand binding.