Application Note
Measuring Mouse GFR By FITC-Inulin Using The Thermo Scientific NanoDrop 3300 Fluorospectrometer
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•Application Note: Measuring Mouse GFR By FITC-Inulin Using The Thermo Scientific NanoDrop 3300 Fluorospectrometer
By Thermo Scientific NanoDrop Products
The measurement of glomerular filtration rate (GFR) is the gold standard in kidney function assessment, and as such is used as an indicator of kidney disease (1). Currently, GFR is determined by measuring the level of the endogenous biomarker creatinine (2) or radioactively (3H or 14C) labeled inulin in blood plasma and/or urine (1). Rodents, particularly mice, have high concentrations of non-creatinine chromagens in plasma and urine that interfere with commercially available creatinine assays. As a consequence, these assays, which are based on the alkaline picrate Jaffé reaction or enzymatic reactions, overestimate creatinine concentration in plasma compared with HPLC termination (2, 3). In addition to this, creatinine has also been proven to be secreted in the proximal tubule of the mouse, further adding to the inaccuracy of creatinine-based estimates of GFR (4). For these reasons, alternate tracer molecules have been sought to help increase the accuracy of GFR measurements.
GFR measurements using radioactive labeled inulin clearance, the traditional gold-standard, typically require approximately 3-5 µl of sample per measurement. This inulin radioassay can be used in both anesthetized and conscious mice, but harbors the disadvantages of numerous safety regulations and strict handling issues inherent with the use of radioactive isotopes.An alternative protocol presented by Qi et al. (5) utilizes FITC-labeled inulin as the exogenous marker to measure GFR in conscious mice. This protocol retains the sensitivity of the traditional assay by using fluorescence detection and circumvents the disadvantages of radiolabel-based assays. The FITC-inulin assay has been used in a reduced volume format to take advantage of the microvolume capability of the Thermo Scientific NanoDrop 3300 Fluorospectrometer in order to further reduce the total volume of the assay (4, 6-12). The NanoDrop™ 3300 uses a patented "cuvette-less" sample retention system for fluorescence measurements using samples as small as 1-2 µl.
Click Here To Download:•Application Note: Measuring Mouse GFR By FITC-Inulin Using The Thermo Scientific NanoDrop 3300 Fluorospectrometer
