Fluorescein isothiocyanate (FITC)–inulin (150 mg) was dissolved in 0.9% NaCl (3 ml) at 75°C and dialyzed in 0.9% NaCl (1000 ml) at 25°C for 24 hours. Dialyzed FITC-inulin (3.74 μl/g body weight) was intravenously injected into living C57BL/6 mice at t = 8, 12, 16, 24, and 48 hours after treatment with CDDP-treated (20 mg/kg body weight) or saline-treated (0.2 ml) mice. Blood (approximately 20 μl) was collected via orbital at 3, 7, 10, 15, 35, 55, and 75 min after injection of FITC-inulin and then centrifuged for 20 min at 3500 rpm. The serum sample (10 μl) was diluted with Hepes buffer (40 μl, 500 mM, pH 7.4), and fluorescence was measured using a SpectraMax with excitation at 485 nm and emission at 538 nm. Serum fluorescence data were presented as a two-component exponential decay curve using nonlinear regression. GFR was calculated according to the equation: GFR = I/(A/α + B/β), where I is the amount of FITC-inulin delivered by the bolus injection, A and B are the y intercept values of the two decay rates, and α and β are the decay constants for the distribution and elimination phases, respectively.
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