MCS papers published in Biosensors and Analytical Biochemistry
In collaboration with the Netherlands Forensic Institute (NFI) MCS published a review paper in the special issue ‘Advances in Lab-on-Chip Devices’ of Biosensors.
Microfluidic devices may offer various advantages for forensic DNA analysis, such as reduced risk of contamination, shorter analysis time and direct application at the crime scene. Microfluidic chip technology has already proven to be functional and effective within medical applications, such as for point-of-care use. In the forensic field, one may expect microfluidic technology to become particularly relevant for the analysis of biological traces containing human DNA. This would require a number of consecutive steps, including sample work up, DNA amplification and detection, as well as secure storage of the sample. The article provides an extensive overview of microfluidic devices for cell lysis, DNA extraction and purification, DNA amplification and detection and analysis techniques for DNA. Topics to be discussed are polymerase chain reaction (PCR) on-chip, digital PCR (dPCR), isothermal amplification on-chip, chip materials, integrated devices and commercially available techniques. A critical overview of the opportunities and challenges of the use of chips is discussed, and developments made in forensic DNA analysis over the past 10–20 years with microfluidic systems are described. Areas in which further research is needed are indicated in a future outlook.
In the top row, the consecutive steps in the process of forensic DNA analysis are listed; within the middle row, the conventional technique with some examples. In the bottom row, microfluidic analogies can be found with some examples.
In this research six cyanine fluorophores for the detection of dsDNA are investigated: EvaGreen, SYBR Green, PicoGreen, AccuClear, AccuBlue NextGen and YOYO-1. For AccuClear and AccuBlue NextGen, dyes which have recently become available on the market, in this paper the first scientific data of their spectral behavior is presented. Measured absorption spectra of all dyes are compared with information of the manufacturer and literature. For each dye the fluorescence intensity as a function of the amount of dsDNA is measured at the optimal wavelengths for excitation and emission. From this, the limits of detection and the response linearities at low levels of dsDNA are determined. For the first time for all six dyes their characteristics are determined under identical conditions and using similar criteria. Additional as well as results striking from literature for some absorption, excitation and emission maxima (for both the free dye and the dye/dsDNA complex) are found, as well as for the linear response of the dyes. No linear range nor limit of detection could be determined for SYBR Green and YOYO-1 for low amounts of dsDNA. EvaGreen, PicoGreen, AccuClear and AccuBlue NextGen show linearity in the pg-ng range. AccuClear exhibits the widest linear range of 3 pg - 200 ng, whereas AccuBlue NextGen turned out to have the highest sensitivity of the tested dyes with a limit of detection of 50 pg.
Fluorescence intensities of the dyes in the presence of 0, 0.1, 0.2, 0.5, 1, 2, 5, 10, 20, 50 & 100 ng salmon dsDNA and 1.0X dye (100 nM for YOYO-1) 200 µL (except for AccuClear where 210 µL is used. Spectra were recorded with a M200 PRO multimode reader (Tecan). The error bars are ± 1 standard deviation. EvaGreen, SYBR Green, PicoGreen and AccuClear are depicted on the left y-axis, whereas YOYO-1 is depicted on the right x-axis, as is indicated by the coloured arrows in the graph.
Link to the article: http://authors.elsevier.com/a/1TYexlbAHrYT