There are currently several absorbance- and fluorescence-based protein assays in common use, each with its own shortcomings, such as protein-to-protein variability, contaminant interference, time requirements, accuracy, sensitivity, and the need for caustic or hazardous reagents. We compared four common protein assays for protein-to-protein variation, accuracy, precision, and sensitivity. The Quant-iT™ Protein Assay for the Qubit® Fluorometer compared favorably by providing low protein variability, rapid quantitation, accuracy, precision, and high sensitivity.
NOTE: Since this article was published, we have launched the next generation instrument, the Qubit® 2.0 Fluorometer which quantitates DNA, RNA, and protein with unprecedented accuracy, sensitivity, and simplicity.
The Qubit™ Quantitation Platform: Fast and Easy to Use
Figure 1. Workflow for the Quant-iT™ assays using the Qubit® Fluorometer.
Quant-iT™ Protein Assay: Accurate, Precise, and Sensitive
The Quant-iT™ Protein Assay is insensitive to many common contaminants, including reducing agents, nucleic acids, and free amino acids. However, detergents, such as SDS (final concentration >0.01%), Tween® 20, and Triton® X-100, are not recommended. The assay has an optimal range of 1.25 to 25 µg/mL (0.25–5 µg) in the assay tube (initial stock concentrations, 12.5–5 mg/mL) and is provided in a simple kit format that allows easy and rapid use. Learn more about the Qubit™ Quantitation Platform.
Figure 2. The Quant-iT™ Protein Assay with the Qubit® Fluorometer produces less protein-to-protein variation and higher accuracy, precision, and sensitivity than 3 other common protein assays. (A–D) The same lot of each protein was used in all assays, and assays were carried out in triplicate following the manufacturers’ protocols. Data are graphed to show protein-to-protein variation throughout the protein concentration range tested. The inset in (A) is a magnification of the low end of the protein range, to show the sensitivity of the Quant-iT™ Protein Assay used with the Qubit® Fluorometer.
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