This method is suitable for materials with a critical diol structure, such as cellulose, agarose, and dextran. Oxidation of periodate results in the formation of an acetaldehyde group, which reacts with the amino group to form a Schiff base and is stabilized by borohydride reduction. Source: Laboratory Manual of Enzymology

Operation method

basic program

Materials and Instruments

Cellulose powder
NaHCO3-NaIO4 Sodium carbonate buffer.NaBH4 solution Sodium acetate-acetic acid PBS

Move

See "Others" for "Reagents" required for the experiment.

Suspend 2 g of cellulose in 40 ml of 0.1 mol/L _NaHCO3-0.06 mol/L _NaIO4 and leave for 2 h under dark condition, then rinse with 0.1 mol/L pH 9.0 sodium carbonate buffer on a glass stock. Proteins to be immobilized (~1 mg/ml) were added to 40 ml of buffer and shaken overnight at room temperature. To remove Schiff base, 2 ml _NaBH4 was added to the solution and shaken at room temperature for 30 min, after which 2 ml _NaBH4 was added again and shaken at room temperature for 30 min. The solid material was rinsed with 0.1 mol/L pH 4.0 acetate-acetic acid, PBS and 0.05% Tween 20.

Common Problems

Reagents:

0.1 mol/L _NaHCO3-0.06 mol/L _NaIO4 ( _NaHCO3, Mr = 84.0; _NaIO4, Mr = 213.9; 8.4 g _NaHCO3, 12.8 g _NaHCO3 dissolved in 100 ml _H2O )

0.1 mol/L sodium carbonate buffer (NaHCO3-Na2C03), pH 9.0

_NaBH4 solution (0.1 g _NaBH4 dissolved in 10 ml 0.1 mol/L NaOH, freshly prepared)

0.1 mol/L sodium acetate-acetic acid, pH 4.0

PBS (phosphate buffer: 10 mmol/L potassium-sodium dihydrogen phosphate, pH 7.2, 0.8% NaCl, 0.02% KCl)

0.05% Tween 20 dissolved in PBS

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