Automated, quantitative DNA purification and spectrometric analysis from whole blood samples in a compartmented reactor device

Abstract

The purification of nucleic acids plays a major role in the fields of biotechnology and medicine. In this study, the systematic optimization of an automated, multi-step sample-to-answer purification process of genomic DNA starting from human whole blood samples is reported. The process uses an in-house developed, robust reactor device including microfluidic segmented flow, contactless magnetic particle separation and resuspension as well as spectrometric analysis for quality control and quantification. At first, the integrated quality control was validated using external methods like agarose gel electrophoresis, SDS-PAGE and, in order to examine integrity and functionality of the purified DNA, PCR. The device was able to achieve comparable DNA yields in identical quality compared to manually conducted control experiments (average DNA eluate concentrations 16.6 ± 1.1 μg mL⁻¹versus 17.5 ± 2.9 μg mL⁻¹). Furthermore, electromagnetic resuspension, the amount of magnetic particles used for a purification experiment as well as the number of conducted wash steps have been identified as major parameters influencing process performance. Concluding, a cleaning-in-place procedure was developed and evaluated, indicating the elimination of any DNA residues between successive purification processes and thereby potentially qualifying the reactor device for diagnostic purposes.