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Comparative study of thiophilic functionalised matrices for polyclonal F(ab')2 purification.
Peter Kumpalume1, Nigel K H Slater
1Department of Chemical Engineering, University of Cambridge, Pembroke Street, Cambridge CB2 3RA, UK.
Journal of Chromatography. A
|February 3, 2004
Summary
New thiophilic adsorbents demonstrate high selectivity and capacity for binding F(ab')2 fragments. Thiol-immobilized ligands show superior performance compared to amino-immobilized ones for antibody purification.
Area of Science:
- Biochemistry
- Protein Chemistry
- Chromatography
Background:
- Antibody fragments are crucial in diagnostics and therapeutics.
- Efficient purification methods are needed for F(ab")2 fragments.
- Thiophilic adsorbents offer potential for selective protein binding.
Purpose of the Study:
- To develop and evaluate thiophilic adsorbents for F(ab")2 fragment purification.
- To compare the binding capacity and selectivity of different immobilized ligands.
- To assess the performance of novel adsorbents for antibody fragment recovery.
Main Methods:
- Development of thiophilic adsorbents using divinyl sulfone or epoxy activated Streamline quartz.
- Testing capacity and selectivity for polyclonal F(ab")2 fragments.
- Comparison of thiol and amino immobilized ligands.
- Evaluation of dynamic binding capacities and recovery rates.
Main Results:
- All tested adsorbents, except epoxy activated guanidine, showed high F(ab")2 selectivity.
- Dynamic binding capacities ranged from 3 to 10 mg/ml.
- Thiol-immobilized ligands exhibited higher F(ab")2 adsorption and recovery than amino-immobilized ligands.
- Adsorbents demonstrated good selectivity for IgG with better dynamic binding capacities than for F(ab").
Conclusions:
- Thiophilic adsorbents are effective for purifying F(ab")2 fragments.
- Thiol-immobilized ligands provide enhanced performance for F(ab")2 binding and recovery.
- These adsorbents show promise for large-scale antibody fragment purification.