Principle, operation and calculation of Lowry method for protein determination在线视频

The Lowry protein assay is a biochemical assay

for determining the total level of protein in a solution.

The total protein concentration is exhibited by a color change

of the sample solution in proportion to protein concentration,

which can then be measured using spectrophotometer.

It is named for the biochemist Oliver H. Lowry

who developed the technique in the 1940s.

His 1951 paper describing the technique

is one of the most-highly cited papers ever in the scientific literature,

cited over 200,000 times.

The Lowry method is based on the reaction of cupric ion （Cu+ ）

produced by the oxidation of peptide bonds

with Folin reagent (a mixture of phosphomolybdate and phosphotungstate)

under alkaline conditions.

The reactions result in a strong blue color,

which depends mainly on the amino acids residues,

such as tyrosine and tryptophan,

the intensity of which is related to the content of protein.

In this experiment,

the samples and the standard protein solution are treated.

After color development,

the absorbance of each sample is determined by absorbance at 650 nm,

thus the protein content can be calculated.

Prepare 7 clean test tubes

and number them (1,2,3,4,5,6,7) near the top

for making the standard curve and analyzing the sample.

and proceed according to the table below.

Mix completely,

incubate at 50 degrees celsuis for 10 min ,

then cool to room temperature.

Incubate at room temperature for 10 min

Mix immediately,

incubate at 50 degrees celsuis for 10 min, then cool ,

Set the absorbance of tube 6 at 650 nm to zero.

Read and record the absorbances of other tubes.

Fill in the table for your raw data

Determine the protein concentration of tubes 7 from your graph.

Calculate the protein concentration of the sample

according to the following formula