This technique is more efficient and gentle compared to the chloride method. Source: Laboratory Manual of Enzymology
Operation method
basic program
Materials and Instruments
Enzyme solutions Move See "Other" for "Reagents" required for the experiment. 1. Alkalysis and activation Polyester pellets (2 cm2 each, 10 pellets) were incubated in 10 ml of 10% (mass concentration) NaOH for 10 min at room temperature. the pellets were rinsed five times with water, 10 ml each time, and then submerged in dioxane for 30 min, and then rinsed five times with dioxane to remove any residual water. Subsequently, the polyester pellets were incubated in dioxane dissolved in 35 g/l of carbonyldiimidazole for 30 min. The pellets were finally rinsed five times with dioxane and three times with 0.1 mol/L pH 7.5 potassium phosphate. 2. Enzyme coupling The polyester pellets were submerged in 10 ml of enzyme solution and incubated overnight at room temperature or at 4°C for sensitive enzymes. Unbound enzyme was removed by rinsing on a suction filter with 0.1 mol/L pH 7.5 potassium phosphate dissolved in 0.5 mol/L NaCl followed by 0.1 mol/L pH 7.5 potassium phosphate. The polyester pellets can be stored in buffer at 4°C for one week. For longer storage, these polyester chips must be rinsed briefly with water and dried in air (not recommended for use in sensitive enzymes). Common Problems Reagents: 0.1 mol/L potassium phosphate pH 7.5 0.1 mol/L potassium phosphate dissolved in 0.5 mol/L NaCl, pH 7.5 100 g/l NaOH Dioxane Dioxane dissolved in 35 g/l carbonyl diimidazole (CDI, Mr = 162.2) For more product details, please visit Aladdin Scientific website.
Potassium phosphate NaCl NaOH Dioxane Carbonyldiimidazole
Suction filter Polyester tabs