Application Note: Automated Washing Of Magnetic Bead QuantiGene® Plex 2.0 Assays

By Paul Held, Principal Scientist, Applications Dept., BioTek Instruments, Inc.

The characterization of gene expression through the quantitation of mRNA species from experimental samples is a commonly used technique in biomedical research today. The QuantiGene® Plex 2.0 assay kit from Affymetrix offers a means to quantitate multiple mRNA species simultaneously from a variety of different sample types. This technology, which uses ferrite beads as a solid substrate for the reactions requires a magnetic field for bead retention during the wash steps. Here we describe the use of the ELx405™ Magnetic Bead Washer to automate the wash steps of this assay technology.

Quantitating and characterizing gene expression through RNA measurement is a commonly performed task in today's biomedical research. Typically RNA expression is examined through quantitative PCR, which requires extensive RNA purification and amplification. These methods are time consuming, expensive, and prone to errors. To rectify these problems, alternative methods have been developed. One such method is the QuantiGenePlex 2.0 System, which measures RNA directly from tissue homogenates, whole blood, cultured cell lysates or purified RNA without target amplification. The QuantiGene Plex 2.0 System uses branched DNA (bDNA) technology in conjunction with multi-analyte magnetic beads (xMAP® technology from Luminex Corporation) to provide the detection and quantitation of multiple mRNA targets simultaneously. The bDNA technology is a hybridization based methodology that uses labeled DNA probes to amplify the signal rather than the target mRNA. The xMAP technology uses flow cytometry and fluorescently-dyed microspheres (beads) to allow multiplexing up to 500 unique assays within a single sample. Each hybridization step requires a thorough wash step to remove the unhybridized material. In order to increase throughput and precision of the assay, there was a desire to automate the wash steps employed by the assay.