November 15, 2006 (Vol. 26, No. 20)
Promega’s Maxwell® 16 Automates the Purification of Biomolecular Compounds
The rapid and effective preparation of biomolecules is a required first step in many common laboratory preparative and analytical techniques. The use of varying degrees of automation can provide advantages by offering consistency beyond the capabilities of manual preparative methods. Promega (www.promega.com) has developed the Maxwell®16 System to bring personal automation to laboratories that have traditionally not had access to automation platforms.
The system is comprised of integrated instrumentation, prefilled reagent cartridge kits, preinstalled automated methods, and technical support to allow a breadth of applications. Maxwell 16 reagent kits designed for purifying genomic DNA (gDNA), RNA, forensic samples, and polyhistidine-tagged proteins run on a common instrument platform. The System is designed to provide consistent performance, ease-of-use, and ability to run multiple reagent chemistries on the same instrument to meet the changing needs of a research laboratory.
Figure 1
System Design
The Maxwell 16 System consists of a compact, benchtop Maxwell 16 Instrument, automated methods, and reagent kits containing prefilled reagent cartridges. The system was designed to provide for the changing needs of research labs and can be easily upgraded with new capabilities as reagent cartridges and software are released.
The Maxwell 16 Instrument purifies biomolecules through the sequential capture and release of coated paramagnetic particles (PMPs) into the wells of prefilled reagent cartridges (Figure 1). The Maxwell 16 Instrument utilizes powerful capture magnets and unique plunger design to capture the PMPs with attached biomolecules and to wash away impurities during a run. The ability of the instrument to selectively capture and move PMPs during processing eliminates the need for complex liquid handling that can result in clogged filtration columns or contaminated tubing. One prefilled reagent cartridge, plunger, and elution tube are used per purification and are disposable.
The single-use, prefilled reagent cartridges and specialized plastics ensure that reagents are delivered fresh and untainted to maximize performance and consistency. Purified samples are eluted into elution buffer ready for use; precipitation and rehydration is not required. From 1–16 samples can be processed in about 30 minutes.
Figure 2
Purification Principle
Promega developed the MagneSil® purification matrix to combine the proven performance of silica-based purification methods with the advanced handling characteristics inherent in PMPs. The MagneSil PMPs consist of a silicon dioxide:magnetite composition that generates a surface capable of binding nucleic acids and other biomolecules in a format that can be manipulated using a magnetic field. MagneSil PMPs migrate in a magnetic field but retain no magnetic memory, ensuring rapid capture and resuspension. MagneSil PMPs also demonstrate extremely fast magnetic response times and have no iron leach, features that are desirable for the efficient isolation of highly pure biomolecules.
The PMP format can be considered a mobile solid phase that offers scaleable solution-based binding kinetics with the ability to easily wash and elute materials of interest. The design of particles of varying pore size and the ability to alter the binding/wash conditions has generated a family of chemistries that have been successfully used for DNA and RNA purification in research, clinical, and forensic applications. A related MagneHis™ PMPs is used for polyhistidine- and HQ-tagged protein purification. Currently, Maxwell 16 isolation kits are available for gDNA, total RNA, polyhistidine- and HQ-tagged protein and forensic gDNA purification applications.
Figure 3
System Performance
The Maxwell 16 System is designed to allow multiple reagent chemistries to run on a single instrument. The Maxwell 16 gDNA Isolation Kits have been validated for use with whole blood (400 µL), tissue culture cells (5×106), and a variety of whole organism and tissue types (Figure 2a). The unique design of the Maxwell 16 Plunger allows the direct processing of a variety of solid sample types such as whole Drosophila flies, animal and plant tissues, and rodent tails without preprocessing.
Once samples are placed into prefilled reagent cartridges, the cartridges are loaded into the instrument, and an automated method is selected. Purified gDNA is delivered in a concentrated format ready for downstream analysis without the need for differential lysis, fractionation of white cells, centrifugation, precipitation, or rehydration of gDNA prior to analysis. Yield and performance in downstream applications such as PCR are shown in Figure 2b.
The Maxwell 16 Total RNA Isolation Kit can be used with a variety of sample sources and is effective at isolating RNA from samples containing a wide range of total RNA content. For most tissue types up to 50 mg of sample can be processed. The system uses a proprietary clearing agent to effectively remove contaminating gDNA without using a nuclease, and fewer than 0.1 genomic equivalents are present per 100 ng of isolated RNA as determined by qPCR analysis. Most tissue types can be prepared for purification by disruption in RNA Lysis Buffer using mechanical shearing.
We have also isolated total RNA from stabilized blood, the white blood cell fraction of whole blood, and plant leaf tissue. Yields are excellent with A260/A280 ratios of 2.0 or better. The isolated RNA is of high quality, and performance in downstream applications is excellent (Figure 3).
The Maxwell 16 Polyhistidine Protein Purification kit is designed to isolate recombinant polyhistidine- or HQ-tagged proteins from bacterial cells, mammalian cells, insect cells, or tissue culture medium (Figure 4). Sample or lysate is added directly to the reagent cartridge, and purified proteins are obtained in about 40 minutes of hands-free operation. Once purified, proteins are ready for downstream analysis using a variety of techniques such as polyacrylamide gel analysis, Western blot, MALDI-TOF mass spectrometry, and functional analysis.
Figure 4
Conclusion
The Maxwell 16 System is designed to offer consistent performance in an easy-to-use format. The available reagent kits for gDNA, RNA, forensic gDNA, and polyhistidine-tagged protein isolation run on a common compact benchtop instrument platform.
Dan Kephart, Ph.D., Terri Grunst, Steve Krueger, Katharine Hoffman, and Hemanth Shenoi, Ph.D., are with Promega. Web: www.promega.com. Phone: (608) 277-2490. E-mail:hemanth.shenoi@ promega.com.
