Analysis and purification of IgA from Shanghai Xinfan Technology Lecture - Huaqiang

The probe capacitor polarity needle has a head diameter of 5.8 mm. The total length of the 7-pin and 12-pin needles is 5 cm.

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1. DEAE-52 Cellulose

2. 0.10 mol/L ZnSO₄ solution: Dissolve 2.88 g of ZnSO₄·7H₂O in water to make up to 1000.00 ml.

3. Saturated Ammonium Sulfate Solution

4. 10% EDTA-Na Solution

5. Sephadex G-200

6. 0.01 mol/L pH 7.4 PBS Solution

7. 0.10 mol/L pH 6.4 PBS Solution

8. Serum Sample

1. Mix serum with an equal volume of 0.1 mol/L ZnSO₄ solution, adjust pH to 7.0, stir at room temperature for 1 hour, then centrifuge to collect the precipitate.

2. In the supernatant, add an equal volume of saturated ammonium sulfate solution, mix well, and let it stand for 30 minutes or overnight in the refrigerator.

3. Centrifuge at 10,000 rpm for 10 minutes, and discard the supernatant.

4. Dissolve the precipitate in 4 ml of 10% EDTA-Na solution.

5. Perform dialysis in normal saline to remove impurities.

6. Load the sample onto a DEAE-52 column using 0.01 mol/L pH 7.4 PBS buffer. Elute the protein (IgG) and discard it.

7. Further elute IgA using 0.1 mol/L pH 6.4 PBS buffer.

8. Pass the collected fraction through a Sephadex G-200 column using 0.01 mol/L pH 7.4 PBS buffer (0.14 mol/L NaCl). Collect the eluent and measure the OD₂₈₀ value. The first peak corresponds to purified IgA.

9. Perform final dialysis and concentration for further analysis and identification.