The equilibration step is adding the serum sample to the buffer-equilibrated (pH 6) magnetic Protein G beads. What was expected to occur if this step was missed? When we mix the samples on the mixer, it was crucial that there was no foam present in the sample - why is this and what part of the mixture would be "foamy" at this step? Lastly, we use an neutralizing buffer after eluting the samples from the column. Why is this step important?
Recombinant Protein G from Streptococcus are covalently attached in an oriented fashion to magnetic beads. Bovine Serum is added and the antibodies are captured by the beads. Using the magnetic device, beads are attached and unbound material is washed away. Antibodies are eluted using a lower pH buffer and the solution is neutralized.
The equilibration step is adding the serum sample to the buffer-equilibrated (pH 6) magnetic Protein G beads. What was expected to occur if this step was missed? When we mix the samples on the mixer, it was crucial that there was no foam present in the sample - why is this and what part of the mixture would be "foamy" at this step? Lastly, we use an neutralizing buffer after eluting the samples from the column. Why is this step important?
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