< 0.5% cross-reactivity observed with available related molecules.< 50% cross-species reactivity observed with species tested.
No significant interference observed with available related molecules.
The Quantikine Mouse/Rat FGF-21 immunoassay is a 4.5 hour solid-phase ELISA designed to measure FGF-21 in mouse or rat cell culture supernates, serum, and plasma. It contains E. coli-expressed recombinant mouse FGF-21 and antibodies raised against the recombinant factor. This immunoassay has been shown to accurately quantitate the recombinant mouse FGF-21. Results obtained using natural mouse or rat FGF-21 showed dose response curves that were parallel to the standard curves obtained using the Quantikine mouse kit standards. These results indicate that this kit can be used to determine relative mass values for natural FGF-21.
Intra-Assay Precision (Precision within an assay) Three samples of known concentration were tested in separate assays to assess inter-assay precision.
Inter-Assay Precision (Precision between assays Three samples of known concentration were tested on one plate to assess intra-assay precision.
Cell Culture Supernates
Serum, EDTA Plasma, Heparin Plasma
The recovery of FGF-21 spiked to levels throughout the range of the assay in various matrices was evaluated.
Average % Recovery
Mouse Cell Culture Samples (n=6)
Mouse EDTA Plasma (n=4)
Mouse Heparin Plasma (n=4)
Mouse Serum (n=4)
Rat Cell Culture Samples (n=2)
Rat EDTA Plasma (n=4)
Rat Heparin Plasma (n=4)
Rat Serum (n=4)
To assess the linearity of the assay, samples spiked with high concentrations of FGF-21 in each matrix were diluted with the appropriate Calibrator Diluent and assayed.
FGF-21 is a 209 aa polypeptide that presumably has a signal sequence and is secreted. Human FGF-21 demonstrates only 75% aa identity with mouse FGF-21. In mouse, it is expressed preferentially in liver.
Refer to the product for complete assay procedure.
Bring all reagents and samples to room temperature before use. It is recommended that all samples, standards, and controls be assayed in duplicate.
Prepare all reagents, standard dilutions, and samples as directed in the product insert.
Remove excess microplate strips from the plate frame, return them to the foil pouch containing the desiccant pack, and reseal.
50 µL Assay Diluent
Add 50 µL of Assay Diluent to each well.
50 µL Standard, Control, or Sample
Add 50 µL of Standard, Control, or sample to each well. Cover with a plate sealer, and incubate at room temperature for 2 hours.
Aspirate each well and wash, repeating the process 4 times for a total of 5 washes.
100 µL Conjugate
Add 100 µL of Conjugate to each well. Cover with a new plate sealer, and incubate at room temperature for 2 hours.
Aspirate and wash 5 times.
100 µL Substrate Solution
Add 100 µL Substrate Solution to each well. Incubate at room temperature for 30 minutes. PROTECT FROM LIGHT.
100 µL Stop Solution
Add 100 µL of Stop Solution to each well. Read at 450 nm within 30 minutes. Set wavelength correction to 540 nm or 570 nm.
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