Folate measurement in mammalian tissues by fluorescence polarization

Abstract

Measurement of folate in animal tissues is expensive and time-consuming. A fluorescence polarization assay has been developed that allows the rapid and inexpensive quantification of folate in various animal tissues. The concentration of tissue folate is determined by its ability to compete with the binding of Alexa-660-folate to bovine milk folate binding protein. The technique uses a few milligrams of material and is amenable to automated screening in 384-well microplates. Using this approach, the folate concentration in mouse liver, kidney & brain was found to be 21.4, 4.22 and 0.73 nanomoles/g fresh tissue, respectively. Packed human erythrocytes were found to contain 1.31 mM folate. These estimates are similar to published folate values for these tissues. Ascorbate was not included in the assay buffer because of its pro-oxidant effects in iron rich tissues such as erythrocytes and liver. The assay is homogeneous, completed within a few hours of the availability of the samples, and will enable the high throughput analyses of folate in human and animal samples.