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Article
April 1982

Measurement of Fluorescein Binding in Human Plasma Using Fluorescence Polarization

Author Affiliations

From the Departments of Ophthalmology (Drs Brubaker and Nagataki and Mr Grotte) and Cell Biology (Dr Penniston), Mayo Clinic, Rochester, Minn. Dr Nagataki is a Research to Prevent Blindness International Research Scholar.

Arch Ophthalmol. 1982;100(4):625-630. doi:10.1001/archopht.1982.01030030627020
Abstract

• The binding of fluorescein was measured in human plasma using four methods: steady-state dialysis, pressure dialysis, equilibrium dialysis, and fluorescence polarization. All methods indicated that fluorescein is approximately 85% bound throughout a wide range of concentrations. Technical difficulties were encountered with the fluorescence polarization method above fluorescein concentrations of 10-5 g/mL, but below this concentration, the method gave reproducible and reliable results compared with the three other methods. The number of binding sites available in plasma was estimated to be 4 x 10-3M, and the dissociation constant was estimated to be 6 x 10-4M. These estimates imply that the boundfree ratio of fluorescein in plasma is almost constant at concentrations below 10-4 g/mL. Fluorescein binding can affect the fluorescent intensity or the rate at which fluorescein penetrates the blood-ocular barriers.

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