Parallel Capillary Electrophoresis:
Description of Technology

Advanced Analytical's parallel capillary electrophoresis (CE) technology enables medium to large scale electro driven analytical separations of small molecules and biomolecules with on-line, simultaneous UV absorbance detection. Systems are comprised of an exchangeable capillary array assembly, a fixed optical detection platform, a high voltage power supply, a fluid pumping system, and an automatic tray positioning system. Customized software is tailored for specific applications and provides instrument control, data acquisition, and data analysis.

Two different platforms have been developed: the 96XT series houses 96 capillaries and the newly developed 24HT series houses 24 capillaries. The standard capillary dimensions are 50μm or 75μm inner diameter (i.d.) and 200μm outer diameter (o.d.). Various capillary lengths are used, depending on the application, required resolution and desired speed of analysis. The inlet side of the capillary array is arranged in an 8 x 12 format with electrodes placed side-by-side with each capillary, allowing for direct injection via vacuum or voltage from standard 96-well microtiter plates. The capillaries are aligned in parallel in the detection window, which is positioned between the UV light source and the detector assembly. The outlets of the capillaries are bundled to a common reservoir where they are connected to a ground. The capillary arrays are modular in nature and components can be exchanged in less than five minutes with no tools required.

Room temperature air is circulated around the capillaries to modulate the temperature during the CE separation. Alternatively, a Peltier cooler is available to actively cool the capillary array for performing higher current applications such as log P or chiral analysis.

Technology Schematic (96XT Version shown)

The optical platform consists of a UV light source, collimating lens, UV camera lens and a 1024-element linear photo diode array (PDA) detector. The UV light is provided by a line source consisting of a zinc (214 nm), cadmium (229 nm), or mercury (254 nm) lamp. A broadband deuterium source may also be used. Detection is performed at a fixed, single wavelength determined by the UV lamp and an interference filter. Light from the lamp passes through a collimating lens, the detection windows of the capillaries, a focusing UV camera lens, and is then spatially imaged onto the PDA. The capillary array is imaged continuously, providing real time, on-line UV detection for either 24 or 96 different CE separations.

The 20 kV, 6 mA total current high voltage power supply provides the appropriate voltage for electrokinetic sample injection and electrophoresis. The recommended operating limits are +/-16 kV, 3 mA total current (30μA per capillary). Positive or negative polarity power supplies are available and can be easily interchanged.

A syringe pump provides controlled pressure or vacuum to the outlet reservoir of the capillary array. Vacuum can be used for vacuum injection of sample, for introducing buffer solutions into the capillary array from the inlet tray, or for performing vacuum-assisted electrophoresis. A distribution valve allows up to five different buffer bottles and a waste bottle to be simultaneously connected to the system. Systems configured for DNA and RNA oligonucleotide analysis utilize a high-pressure pump in place of the syringe pump to account for the increased viscosity of gel separation matrices.

Automated Tray Positioning System

The automatic tray positioning system provides space for up to four 96-well plates. The four positions on the stage are typically assigned to a waste tray, the inlet run buffer tray, and two sample trays, respectively. The tray handler slides out exterior to the system, allowing for optional interface to a robotic arm device for performing long term unattended operation.

Custom software provides complete automation for performing capillary conditioning, sample injection, electrophoresis separation and data processing. Five different analyzer systems have been developed to date utilizing this technology platform:

• pKa PRO™ 96XT and pKa PRO™ 24HT systems are dedicated, application specific systems for performing physicochemical profiling of drug compounds (pKa, Log P) or chiral separations.
• Oligo PRO™ 96XT and Oligo PRO™ 24HT systems are dedicated, application specific systems for measuring DNA and RNA oligonucleotide or RNAi duplex purity.
• The cePRO 9600™ system is a general purpose version of the platform.

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