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Product Name: Alkali-tolerant Protein A Sepharose FF (rat-PA Sepharose FF)
Catalog Number: NRPB06L, NRPB06S (prepacked column)
Packing Details: 20 ml, 100 ml, 1 L, 1 ml (prepacked column),5ml (prepacked column), 10ml (prepacked column)
Introduction:
Protein A is a cell wall protein of Staphylococcus aureus and contains five functional domains that can bind to the Fc fragment of IgG. Structural modification of nature protein A improved its alkaline resistance. Therefore, NaOH could be used to regenerate resin. A design that   spacer arms on sepharose backbone are conjugated with the oriented binding sites of recombinant alkali-tolerant protein A ensures a formation of strong and efficient conjugate between sepharose and recombinant alkali-tolerant protein A as well as a higher capacity  of recombinant alkali-tolerant protein A than native protein A.
Column Characteristics:
Support
6% highly cross-linked spherical agarose
Ligand
rat-PA
Ligand Density (mg/ml)
>6
Particle Size (レm)
45~165
Flow Rate-Recommended/Maximum (cm/h)
100/750
pH Limits-Working/Cleaning
3~11/2~13
Maximum Operating Pressure (MPa)
0.3
Capacity
∶35 mg/ml IgG
Storage
4PP oPPC to 8PP oPPC in 20% ethanol
Performing A Separation:
Binding buffer: 20 mM sodium phosphate, 150 mM NaCl, pH 7.4
Elution buffer: 100 mM sodium citrate, pH 4.0
Neutralization buffer: 1M Tris-HCl, pH 9.0
1)         Wash the prepacked 1 ml column with 5~10 column volumes of distilled water to remove 20% ethanol.
2)         Equilibrate the column with 5~10 column volumes of binding buffer.
3)         Dilute 5 ml rabbit serum with binding buffer to 50 ml. Filtrate the diluted serum through a 0.45 レm filter and load the sample.
4)         Wash with 10 column volumes of binding buffer.
5)         Elute with 5 column volumes of elution buffer and neutralize collect fractions with neutralization buffer.
6)         After each separation cycle, regenerate the resin by washing with approximately 3~5 column volumes of 0.1 M NaOH (pH 3.0).
7)Confirm the purity of the collected antibody by SDS-PAGE analysis (>95% purity).

Notice:
1)         It is easy and simple to use the prepacked column. It is not necessary to need any equipment to purify antibody.
2)         Be sure that the antibody is stable under the elution conditions selected. If there is any doubt, titrate the antibody fraction to neutrality immediately upon elution in order not to lose biological activity. To accomplish this, prefill the fraction vessels to 5~10% of the intended fraction volume with a buffer concentrate at about pH7.5 (for example 1 M Tris-HCl or 1 M sodium phosphate).
3)         Make sure temperature of the equilibration buffer identical to that of the packed column to avoid air bubbles. If air has entered the column, the column should be repacked.
4)         It is recommended to regenerate the resin once after 5 times separation cycle in order to extend the service life. Wash the resin with 5 column volumes of 0.1 NaOH and 5~10 column volumes of 70% ethanol sequentially to remove hydrophobic substances.
5)         For longer periods of storage, keep at 4~8ìC in 20% ethanol.
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