Objective: to purify the IgGs from a NF155 positive patient serum.
Sample: J.C. de Haro Mateo (18-851 ICQ)
Separation protocol: Melon Gel IgG Spin Purification Kit
- Equilibrate the Melon Gel IgG Purification Support and Purification Buffer to room temperature (~15 minutes).
- Swirl bottle containing the Purification Support (do not vortex) to obtain an even suspension. To ensure proper gel slurry dispensing, use a wide orifice or cut pipette tip to dispense 500µL of slurry into a Spin Column placed in a microcentrifuge tube.
- Centrifuge the uncapped column/tube assembly for 1 minute at 5000g, then remove the spin column and discard flow-through.
- Add 300µL of Purification Buffer to the column, centrifuge at 5000g for 10 seconds and discard flow-through. Repeat this wash once. Place the bottom cap on the column.
- Dilute serum 1/10 with Pufification Buffer.
- Add 100-500µL of diluted serum to the column. Cap column and incubate for 5 minutes at room temperature.
- Remove bottom cap from the column, loosen top cap and re-insert spin column in the collection tube.
- Centrifuge for 1 minute at 5000g to collect the purified antibody in the microcentrifuge tube.
Concentration protocol: Amicon Ultra-4 (Millipore)
- Centrifuge the sample for 2 minutes at 2000g and 4ºC
