Biology:Standard curve

An example of a standard curve showing the absorbance of different concentrations of protein (two trials for each measurement). The mass of protein in a different sample is determined by determining where on the standard curve it should go - in this case, 30 milligrams.

Multiple samples with known properties are measured and graphed, which then allows the same properties to be determined for unknown samples by interpolation on the graph. The samples with known properties are the standards, and the graph is the standard curve. The concentration of the unknown may be calculated from the mass in the assay.

Example

Samples treated with the Bradford assay. The brown sample (lower absorbance) contains no protein, while the blue sample (higher absorbance) contains protein. The amount of protein in the second sample can be determined by comparison to a standard curve.

The Bradford assay is a colorimetric assay that measures protein concentration. The reagent Coomassie brilliant blue turns blue when it binds to arginine and aromatic amino acids present in proteins, thus increasing the absorbance of the sample. The absorbance is measured using a spectrophotometer, at the maximum absorbance frequency (A_max) of the blue dye (which is 595 nm). In this case, the greater the absorbance, the higher the protein concentration.

Data for known concentrations of protein are used to make the standard curve, plotting concentration on the X axis, and the assay measurement on the Y axis. The same assay is then performed with samples of unknown concentration. To analyze the data, one locates the measurement on the Y-axis that corresponds to the assay measurement of the unknown substance and follows a line to intersect the standard curve. The corresponding value on the X-axis is the concentration of substance in the unknown sample.

See also

• Color
• Linear regression
• Logarithmic scale
• Protein
• Serial dilution

References

• Bradford Assay Background (pdf format)

"Bradford Protein Assay". Bio-Rad Quick Start Bradford Protein Assay Instructional Manual. http://www.bio-rad.com/cmc_upload/Literature/55685/4110065A.pdf. Retrieved May 31, 2005.