All compounds containing two formamide groups (-CONH2-) in the molecule can interact with alkaline copper solution to form a purplish-red compound, this reaction is called the double urea reaction protein molecules have many peptide Jian (-CONH2-) can be this reaction, and the various plasma proteins show the same degree of coloration, but also the same degree of coloration. And the degree of coloration of various plasma proteins is basically the same. This experiment is from Mudanjiang Medical College undergraduate 5-year laboratory guide for testing majors.

Operation method

Experimental determination of total serum proteins by bis-urea method

Principle

All compounds containing two formamide groups (-CONH2-) in the molecule can interact with alkaline copper solution to form a purplish-red compound, this reaction is called the double urea reaction protein molecules have many peptide Jian (-CONH2-) can be this reaction, and the various plasma proteins show the same degree of coloration, but also the same degree of coloration. And all plasma proteins have the same degree of coloration.

Move

I. Experimental operations: according to the table

Mixed, placed in 25 ℃ 30min. (37 ℃ 10min) at a wavelength of 540nm. Cuvette diameter 1.0cm to blank tube zero, read the optical density of each tube.

Calculation

Total serum protein g / L = measurement tube (or corrected) absorbance / standard tube absorbance × protein standard concentration (g / L)

Caveat

1, serum protein content is generally expressed in g / L, because the molecular weight of various proteins is different, can not be expressed in mol / L.

2， phenolphthalein, sodium bromosulfophthalein in alkaline solution color affects the results of bioconjugate determination, dextrose anhydride can be determined by the turbidity of the tube also affects the results of the theory of these interferences can be used in addition to the corresponding standard blank tube, such as the standard blank tube absorbance is too high, can affect the accuracy of the determination.

3， containing lipids very much of the serum, the color should be turbid, available 3.l acetic acid extraction and then colorimetric.Reference value: 60-80g / L

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