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Studying protein aggregation by programmed flow field-flow fractionation using ceramic hollow fibers
Ruohua Zhu1, Wijbren Frankema, Yuli Huo
1Polymer-Analysis Group, van't Hoff Institute for Molecular Sciences, University of Amsterdam, Nieuwe Achtergracht 166, 1018 WV Amsterdam, The Netherlands.
Analytical Chemistry
|July 15, 2005
Summary
Ceramic hollow fibers enable precise protein diffusion coefficient measurements in flow field-flow fractionation. This robust system accurately characterizes protein aggregation, even for large structures, with excellent recovery and durability.
Area of Science:
- Materials Science
- Analytical Chemistry
- Biophysics
Background:
- Ceramic hollow fibers offer potential as separation channels in flow field-flow fractionation (FFF).
- An ultrafiltration membrane layer on alpha-alumina fibers was investigated for its separation capabilities.
Purpose of the Study:
- To evaluate ceramic hollow fibers as semipermeable membranes for FFF.
- To determine the diffusion coefficients of standard proteins and study heat-induced protein aggregation.
Main Methods:
- Utilized alpha-alumina hollow fibers with an inner gamma-alumina layer as FFF separation channels.
- Measured diffusion coefficients of proteins and characterized beta-lactoglobulin (beta-LG) aggregation using multiangle light scattering detection.
Main Results:
- Achieved complete protein recovery and high measurement precision (5-10%) for diffusion coefficients.
- Demonstrated extended sample component size range using programmed cross-flow.
- Observed increased size and extent of beta-LG aggregates with protein concentration and heating time, forming large structures (>100 million Da) in salt solutions.
Conclusions:
- Ceramic hollow fibers are effective and durable for FFF applications.
- The system accurately quantifies protein aggregation, revealing aggregate behavior consistent with flexible chains.