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Byzen Pro® : Temperature Responsive Protein A

Innovation in Antibody Purification

Byzen Pro® is an affinity chromatography resin in which a novel Protein A mutant is used as a ligand. The product provides a new antibody purification scheme where antibody is eluted under neutral pH by changing temperature, avoiding risk of aggregation of antibody.

Human antibody is purified from cell culture by using Protein A affinity chromatography. Conventional Protein A resin uses strongly acidic buffer to elute antibody, exposing the target antibody to risk of aggregation and decreased activity. In contrast, our temperature-responding Protein A (Byzen Pro®) allows antibody to be eluted under neutral pH by simply changing temperature. This eliminates any risk of aggregation and decreased activity of antibody, improving manufacturing efficiency of antibody drugs.

Features

  • Antibody is eluted under neutral pH.
    Byzen Pro® needs no acidic conditions. No risk of aggregation of antibody.
  • Binding and elution are controlled by temperature.
    Antibody is bound below 10°C and eluted between 37 to 40°C.
  • A recovery ratio is 95% or more.

Purification with a closed column

Column Volume 1.0 ml
Bed Height 2.5 cm
Buffer 20 mM Phosphate, 150 mM NaCl, pH 8.0
Flow Rate 1.0 ml/min
Binding Temperature 4°C
Elution Temperature 40°C
Sample Human IgG (1 mg) + Cell lysate (1 ml)

Caution:
Column pressure should stay below 0.3 Mpa when operating chromatography equipment or a pump.

Procedure

  1. Attach a 5 ml loop before the column, and wash the column with 5 CV of H2O.
  2. Immerse the column and the loop into a 4°C water bath and then equilibrate the column with buffer.
  3. Wait for 5 min for temperature equilibration, and then inject a sample.
  4. After wash the column to remove impurities, stop the pump. Take the column and the loop out of the 4oC water bath and immerse them into a 40°C water bath.
  5. Wait for 5 min for temperature equilibration, and then start the pump to elute antibody.

Purification with open column (no jacket)

Purification may be performed by placing the column in a cold room/refrigerator or an incubator set at desired temperature. It is recommended to calibrate a time required to equilibrate the column before use.
When a flow rate by gravity is too slow, use an aspirator, a pump or a syringe to withdraw buffer from the column.

Buffer 20 mM Phosphate, 150 mM NaCl, pH 8.0
Flow Rate Gravity
Binding Temperature 4°C
Elution Temperature 40°C

Procedure

  1. Pack the column with Byzen Pro® using a pipette (a bed height of 2 cm or more is recommended when the concentration of antibody is low).
  2. Steps 3) and 4) should be performed in a cold room or by placing the column in a refrigerator set at 4°C.
  3. Equilibrate the column with 5 CV of buffer by gravity (buffer should be pre-cooled at 4°C).
  4. Make sure that the column temperature is 4°C, and gently apply a sample onto the surface of the resin to allow the sample to flow by gravity.
  5. Apply a 4°C buffer onto the surface of the resin without disturbing the surface to wash the column. Repeat this step until all impurities are removed.
  6. Steps 7) should be performed by placing the column in an incubator set at 40°C.
  7. Make sure that the column temperature is 40°C, and gently apply a 40°C buffer onto the surface of the resin without disturbing the surface to elute antibody by gravity.

Specification

Bead Structure Cross-linked polyvinyl alcohol
Mean Particle Size 70 µm
Max. Back Pressure 0.3 MPa, 3 bar
Flow Rate (recommend) 125 to 250 cm/h
pH Range (recommend) pH 6 to pH 8
Binding Temperature 4°C
Eluting Temperature 37 to 40°C
Dynamic Binding Capacity 26 mg/ml (250 cm/h)
(20 cm column) 34 mg/ml (125 cm/h)
Storage Condition 20% ethanol at 4°C

Storage

  1. Wash the column with 5 CV of H2O.
  2. For a closed column, replace water with 20% ethanol and store at 4°C.
  3. For an open column, replace water with 20% ethanol. Take the resin out of the column and store in a closed container at 4°C.

Specificity

Human IgG1, IgG2 and IgG4
Rabbit IgG
Goat IgG

Ordering Information

Product Cat.No. Storage Quantity Price(US$)  
Byzen Pro® 0809N05 4°C 5 ml 800.00 Buy
Byzen Pro® 0809N25 4°C 25 ml 2,700.00 Buy