PubMed · 4648536
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D R Smith. 1972. Computer linkage for Technicon SMA 4A.. https://doi.org/10.1136/jcp.25.11.1012-b
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High-throughput DNA sequencing techniques are under rapid development currently, mainly triggered by the Human Genome Project. At the present time, slab gel based automated DNA sequencing is the standard procedure, utilizing fluorophore labeling and laser-induced fluorescence detection with scanning technology. In this paper, a novel, fiber-optic bundle based detection system is introduced, where a central illuminating fiber is used for the excitation of the electrophoretically separated fluorophore-labeled DNA sequencing fragments, along with several collecting fibers disposed around the illuminating fiber to collect the emitted fluorescent signal. As a model system, Cy5-labeled DNA sequencing fragments were separated on an ultrathin polyacrylamide slab gel and detected by the fiber bundle based laser-induced fluorescence detection system. A 640-nm diode laser was used to generate the illumination beam, and the emitted light collected by the fiber bundle was detected by a solid-state avalanche photodiode.
We demonstrate an integrated on-line system with a fused-silica capillary as the microreactor for PCR and capillary gel electrophoresis with laser-induced fluorescence detection for DNA typing and disease diagnosis. Two applications have been investigated: the four short tandem repeat (STR) loci vWA, THO1, TPOX and CSF1PO (CTTv) for DNA typing, and DNA probe for human immunodeficiency virus (HIV-1) diagnosis. The CTTv are important loci in forensic and genetic linkage analysis. The PCR technique is a powerful tool in HIV research because it can detect the presence of the virus before any antibody response in the infected person. Thus it is important for early diagnosis. Multiplexed PCR in a fused-silica capillary, on-line injection, DNA denaturation and calibration based on a standard ladder have been successfully combined. Also, on-line liquid flow management, DNA separation and detection have been completely integrated.
The availability of automated, rapid and reliable methods for the systematic toxicological analysis (STA) of drugs and poisons in biosamples is of great importance in clinical and forensic toxicology laboratories. Gas chromatography-continuous scan mass spectrometry (GC-MS) possesses a high potential in STA because of its selectivity and identification power. However, in order to develop a fully automated STA method based on GC-MS two main obstacles have to be overcome: (a) sample preparation is rather sophisticated owing to the need to isolate analytes from the aqueous matrix and to allow a correct GC repartition of polar analytes; (b) the large amount of information collected within a single analysis makes it difficult to isolate relevant analytical information (mass spectra of analytes) from the chemical noise. Using a bench-top GC-MS system equipped with a laboratory robot for sample preparation (the Hewlett-Packard 7686 PrepStation) and an original method for mass spectral purification, a fully automated STA procedure was developed involving isolation of drugs from the sample (whole blood with minimal pretreatment, plasma, urine) by means of solid-phase extraction, derivatization (trimethylsilylation) of the acidic-neutral and of the basic extracts, GC-MS analysis, processing of data, and reporting of results. Each step of the procedure, and the method for data analysis in particular, can be easily integrated with other existing STA methods based on GC-MS.