Lepta rProtein A

Lepta rProtein A is an affinity chromatography resin for the purification of monoclonal and polyclonal antibodies. The recombinant protein A ligand is produced in E. coli and engineered for directional conjugation, which enhances binding capacity while preserving specific binding to the IgG Fc fragment. Epoxy-based coupling ensures low ligand shedding.

Method of use

Chromatographic conditions

Buffer selection. Equilibration, binding and rinse buffer: 20 mM sodium phosphate, 150 mM NaCl, pH 7.0. Elution buffer: 50 mM sodium citrate, pH 3.0.

Flow rate. Select a linear flow rate of 90–500 cm/h according to the column bed height.

Sample pretreatment. Filter the sample through a 0.45 µm microporous membrane before loading to prevent the column from clogging. Adjust the pH and conductivity of the sample to be consistent with the equilibration buffer, using dilution, ultrafiltration or desalting.

Chromatography steps

1. Equilibration. Use equilibration buffer to fully equilibrate the column until the pH and conductivity are stable and consistent with the equilibration buffer. This step usually requires 3–5 column bed volumes (CV).
2. Sample loading. Determine the loading volume and loading amount from the binding capacity measured in a small-scale test.
3. Washing. Use equilibration buffer or another suitable buffer to wash the column until the UV signal stabilises and returns to baseline.
4. Elution. Elute at lower pH with the elution buffer.
5. Re-equilibration. Re-equilibrate the column with equilibration buffer.

Cleaning and sterilisation

Contaminants such as lipids, endotoxins and proteins accumulate on the column as the number of uses increases. Determine the frequency of cleaning-in-place (CIP) according to the degree of contamination. Where contamination is considerable, CIP after each use is recommended to ensure repeatability of results and to prolong the working life of the resin.

Recommended cleaning conditions by contaminant type:

• Strongly hydrophobic proteins, lipoproteins and lipids: treat with 0.1% non-ionic detergent at 37°C for 1 minute, then wash with at least 5 CV of binding solution.
• Lipids, alternative method: soak in 70% ethanol for 12 hours, then rinse with at least 5 CV of binding solution.

To reduce microbial load, treat the resin with 20% ethanol solution for a minimum of 6 hours.

Storage

Keep unopened resin in the original container at 2–8°C in a well-ventilated, dry and clean place. Do not freeze. Wash the used column with 2–3 CV of 20% ethanol solution and store at 2–8°C.

Destruction and recycling

Chromatography resin is difficult to degrade in nature, so incineration of waste resin is recommended. For resin that has been in contact with biologically active samples such as viruses and blood, follow local biosafety requirements before destroying or disposing of it.

Packing method

Detailed information on resin packaging is available on request. Please contact your local distributor.

Ordering information

• 652-00025 — 25 ml
• 652-00100 — 100 ml
• 652-00500 — 500 ml
• 652-01000 — 1 L
• 652-05000 — 5 L
• 652-10000 — 10 L
• 652-20000 — 20 L