DOE Human Genome Program Contractor-Grantee Workshop VIII
February 27-March 2, 2000  Santa Fe, NM

27. Fully Automated Multiplexed Capillary Systems for DNA Sample Analysis

Qingbo Li, Thomas E. Kane, Changsheng Liu, Harry Zhao, Gary W. Loge, John Kernan, Songsan Zhou, Kevin Levan, Heidi Monroe, and David Fisk

SpectruMedix Corporation, 2124 Old Gatesburg Road, State College, PA 16803

qbli@spectrumedix.com

SpectruMedix has developed a commercial 96-capillary electrophoresis instrument for DNA analysis. All operation steps are automated, including capillary conditioning, gel filling, sample introduction, electrophoresis, and data acquisition. It can perform seven consecutive runs without human intervention. Simple yet highly efficient optical design renders an extremely robust detection system that shows excellent stability. The instrument uses a CCD detector to simultaneously record fluorescence signal from all 96 capillaries with on-column laser excitation. Multi-wavelength detection is implemented with a miniaturized spectrometer utilizing a transmission grating. A replaceable linear-polymer matrix provides high-performance separation for DNA sequencing and genotyping. Currently, the instrument is capable of routinely separating 500+ bases with 98% basecalling accuracy in a 2-hr run. By using a gel matrix that is optimized for longer read, up to 770 bp separation with 98% basecalling accuracy has been achieved in a 3-hr run including capillary conditioning, gel filling, sample introduction, and electrophoresis. The instrument includes an electronic unit that allows monitoring the electrophoresis current for each of the 96 capillaries. Further, a recently developed algorithm allows automated color deconvolution matrix file construction, avoiding the need for a calibration run. This process further enhances the robustness of the instrument. A comparison of the SCE9600 performance to an ABI377 using the same sample has been performed.

A prototype 384-capillary array electrophoresis instrument has been developed for higher throughput analysis. The 384-capillary instrument design is based on the SCE9600 platform, so the 96-capillary instrument can be readily upgraded to obtain 4X higher throughput. The injection end of the 384-capillary array is configured in 16x24 format so that it is compatible with commercial 384-well microtiter tray technology. The instrument is capable of performing one genotyping or sequencing run within 2 hours. In fully automated mode, the instrument will analyze 4,608 DNA samples in a 24-hour day.

The instrument includes an electronic unit that allows monitoring the electrophoresis current for each of the 96 capillaries. This has proved to be a valuable technique for protocol diagnosis and development. Automated basecalling software analyzes a set of 96-capillary data within minutes. Further, a recently developed algorithm allows automated color deconvolution matrix file construction, avoiding the need for a calibration run. This process further enhances the robustness of the instrument since instrumental drift, if there is any, is automatically corrected by the algorithm. A comparison of the sequencing performance of a SCE9600 to the performance of an ABI377 using the same sample has been performed.

A prototype 384-capillary array electrophoresis instrument has been developed for higher throughput analysis. The 384-capillary instrument design is based on the SCE9600 platform, so the 96-capillary instrument can be readily upgraded to obtain 4X higher throughput. The injection end of the 384-capillary array is configured in 16x24 format so that it is compatible with commercial 384-well microtiter tray technology. The instrument is capable of performing one genotyping or sequencing run within 2 hours. In fully automated mode, the instrument will analyze 4,608 DNA samples in a 24-hour day.