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Improving the Success Rate of Protein Crystallization by Random Microseed Matrix Screening
Published on: August 31, 2013
Semi-automated microseeding of nanolitre crystallization experiments
Thomas S Walter1, Erika J Mancini, Jan Kadlec
1Oxford Protein Production Facility and Division of Structural Biology, Wellcome Trust Centre for Human Genetics, University of Oxford, Roosevelt Drive, Oxford OX3 7BN, England.
Summary
A straightforward semi-automated microseeding technique simplifies nanolitre crystallization experiments. This method enhances high-throughput pipelines by enabling efficient seeding with microcrystals, improving success rates in structural biology research.
Area of Science:
- Structural biology
- Biochemistry
- Crystallography
Background:
- Nanolitre crystallization is crucial for determining protein structures.
- Incorporating microseeds can enhance crystal nucleation and growth.
- High-throughput pipelines require efficient and automatable seeding methods.
Purpose of the Study:
- To describe a simple, semi-automated microseeding procedure for nanolitre crystallization.
- To adapt the procedure for additive-screening and cross-seeding.
- To demonstrate the utility of the method with successful examples.
Main Methods:
- Preparation of a microseed stock solution using microcrystals and a Teflon bead.
- Creation of a dilution series from the microseed stock.
- Dispensing 100 nl droplets into 96-well plates for automated integration.
- Modification for additive-screening and cross-seeding experiments.
Main Results:
- Successful establishment of a semi-automated microseeding protocol.
- Demonstration of microseeding integration into high-throughput crystallization.
- Successful application of modified techniques (additive-screening, cross-seeding) in five cases.
Conclusions:
- The described semi-automated microseeding procedure is effective for nanolitre crystallization.
- The method is adaptable and can be integrated into high-throughput workflows.
- This technique facilitates improved success rates in protein crystallization experiments.
