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January 15, 2012 (Vol. 32, No. 2)

Breaking the Sample-Prep Bottleneck

Automated Lysis Systems Streamline Workflows

  • Sample Lysis Systems

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    Figure 1. An example of the high-performance automated sample-preparation system, FastPrep-24, with associated cryogenic and room temperature sample holder adapters

    The requirements for affordable sample homogenization and lysis equipment include the availability of fast, temperature-controlled processes, with complete homogenization of a variety of sample types, without extensive sample manipulations, and without the possibility of sample cross contamination or sample escape to the surrounding environment.

    Now, more than ever, researchers are requiring parallelism and high throughput, i.e., the ability to process multiple samples simultaneously, and to process samples in varying volumes and sizes, in standard labware formats, and in formats amenable to downstream manual or automatic processing.

    Furthermore, researchers require high reproducibility of results on identical sample types, i.e., that the same process settings produce the same quantity and quality of lysate, with minimum sample to sample variability. In many cases, temperature control is also desired.

    All of these requirements can be conflicting, especially for processing difficult samples because in order to have fast and complete sample homogenization, a significant amount of energy is imparted on the samples and the released macromolecules of interest are exposed to those same milling forces and can be damaged. Therefore, it is necessary to minimize the exposure of released macromolecules by accelerating the sample-homogenization process.

  • Sample Homogenization

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    Typical time and speed settings for a FastPrep-24

    Several automated sample-homogenization systems are currently available. MP Biomedicals manufacturers one such family of systems, including FastPrep-24™ and FastPrep-96™ (Figure 1).

    The FastPrep systems consist of an instrument and a disposable lysing matrix, which includes standard labware ranging from 96-well plates and 2 mL tubes, up to 250 mL bottles, loaded with an optimized mixture of inert lysing matrix particles.

    The lysing matrix beads are application specific, and their composition is optimized to provide fast and quantitative lysis of a selected sample type, in most cases within 40 seconds or less. The lysing beads are made of advanced ceramic materials with different hardness values, sizes, and densities, wherein the larger beads play the role of cascading impactor, and the smaller sharper beads are mainly performing lysis by shearing.

    The FastPrep-24 instrument uses a unique, optimized 3-D motion to disrupt tissues and cells through the multidirectional, simultaneous beating of specialized lysing matrix beads on the sample material. The macroscopic motion of nutation results in an in-tube formation of the strong force fields, which accelerate tube content both alongside the vertical axis of the tubes, as well as in a sideways, angular motion.

    This combination of force fields creates the in-tube environment into which the larger and heavier particles perform grinding homogenization and lysis by cascade impaction, and the smaller sharper particles are taken into the fluid vortex, moving in spiral or circular patterns and performing the shearing of the sample.

    Concurrently, the liquid vortex, as a result of its velocity gradient differentials, performs fluid shearing by vortexing. As there are three simultaneous processes being performed in the tube on a sample, the result is a rapid lysis of even the most difficult samples like bone or plant materials within 40 seconds or less. A table of the most common lysis parameters settings for FastPrep-24 system is provided above.

    The FastPrep system has optional temperature control adapters for cryogenic lysis by passive sample cooling with dry ice. Figure 2 represents RNA extracted from Cassava root, an RNase-rich plant material that is both mechanically difficult to homogenize and represents a special challenge for RNA extraction due to extra high content and activity of the RNase enzymes.

  • Conclusions

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    Figure 2. An example of the RNA extracted out of the Cassava storage roots using the FastPrep-24: samples contain 0.32 µg/µL–110 µg/µL RNA. FastPrep settings: Speed 6.0 for 60 s; Lysing Matrix A with an additional zirconium ball.

    In addition to time savings, the largest benefit of an automated sample lysis system is the increased yield and quality of lysate, which assures consistency between sample preparations and eliminates operator and sample variation. Furthermore, automated lysis systems eliminate the most unpleasant repetitive manual workload and significantly improve safety by lysing potentially bio-hazardous samples within a closed, disposable environment.

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