Nucleic acid amplification technologies (NAATs) offer the most sensitive tests in the clinical laboratory. These techniques are used as a powerful tool for screening and diagnosis of infectious diseases. Isothermal methods, as an alternative to polymerase chain reaction (PCR), require no thermocycling machine and can mostly be performed with reduced time, high throughput, and accurate and reliable results.However, current molecular diagnostic approaches generally need manual analysis by qualified and experienced personal which is a highly complex, time-consuming and labor-intensive task. Thus, the demand for simpler, miniaturized systems and assays for pathogen detection is steadily increasing. Microfluidic platforms and lab-on-a-chip devices have many advantages such as small sample volume, portability and rapid detection time and enable point-of-care diagnosis.In this article, we review several isothermal amplification methods and their implementation in microsystems in relation to quantification of nucleic acids. miniaturized systems can be diverse. The majority of publications are focussed on clinical diagnostics including foodborne organisms for environmental monitoring [10][11][12]. In case of an infectious disease, cultivation and phenotypic characterization are still the standard methods of microbiologists which need days up to weeks to obtain a diagnostic result. Hence, scientists started to make use of nucleic acid amplification methods to accelerate the analysis. The most widespread and well known technique for this purpose is the polymerase chain reaction (PCR) [10,13], which exploits the activity of the DNA polymerase. The method is based on a thermal cycling process consisting of repeated cycles of heating and cooling steps allowing for DNA melting, sequence specific primer binding and enzymatic amplification of the target nucleic acid. The quantitative assessment of target copies is possible by using a real-time PCR approach, where a concentration dependent signal (e.g. fluorescence) increases over time [14]. The latter enables the user to easily assess the pathogen load of patient samples [15][16][17][18]. Since the first example of a PCR on a miniaturized system in 1994 [19,20], numerous devices were presented, which integrate not just the amplification, but also sample preparation and detection steps [9,[21][22][23][24][25][26][27][28]. Just a few of those microsystems have reached the market so far, but some have shown a fascinating potential to replace the classical laboratory-oriented state-of-the art [29][30][31][32][33]. setups has been shown to obtain a similar result [34]. A smaller heat capacity allows for rapid changes in temperature beneficial for the PCR time as well as a higher parallelism of multiple genetic samples [34].However, a major drawback of the integration of PCR to microsystems is the necessity of sophisticated instrumentation. The reaction requires a thermal cycling instrumentation, space and considerable expertise [35]. Therefore, isothermal reactions for the ta...
BackgroundA number of pathogens can cause severe destruction of the periodontal apparatus during the course of periodontitis. The aim of this work was the development of a diagnostic device for the use at the point-of-need for the detection of periodontal pathogens to enable a personalized therapy for treatment of periodontitis.MethodsThis test system is based on the polymerase chain reaction of DNA isolated from periodontal pathogens and was examined to precisely detect species-specific sequences on a rotating chip with lyophilized reagents for polymerase chain reaction. The preservation of the reagents was optimized to ensure their stability during the storage.ResultsIn the current work, we have developed a model point-of-care device and showed a proof of concept. It requires low sample volume, is timesaving and can therefore facilitate early diagnosis and treatment of periodontal diseases.ConclusionsThe developed device can provide fast diagnosis of the composition and amount of patients’ oral flora and might help to assess the stage of periodontitis infection. This can facilitate an optimization of therapeutic approaches in order to prevent some of the more serious consequences of the disease.Electronic supplementary materialThe online version of this article (doi:10.1186/s12903-015-0155-y) contains supplementary material, which is available to authorized users.
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