Pomocí molekulárně genetických metod lze prokázat infekční organismy virového, bakteriálního a houbového původu, stejně jako protozoa a parazitické červy. Molekulární metody detekují specifické úseky v sekvencích nukleových kyselin infekčních agens, a není tedy nutné zachování viability hledaných mikroorganismů. Proto je možné použít tyto metody i pro přímý průkaz infekčního agens z fixované tkáně, nejčastějšího dostupného materiálu v patologii. Tento krátký přehledový článek vychází z více než dvacetiletého fungování molekulárně mikrobiologického úseku v rámci patologie a naším cílem je přiblížit možnosti molekulárně genetické detekce infekčních organismů pro patologickou diagnostiku.
Using molecular methods, infectious organisms of viral, bacterial and fungal origin, as well as protozoa and helminths, can be detected. Molecular methods detect specific segments in the nucleic acid sequences of infectious agents and therefore do not require the maintenance of viability of the microorganisms of interest. Therefore, these methods can also be used for direct detection of infectious agents from fixed tissue, the most commonly available material in pathology. This short review article is based on more than 20 years of molecular microbiology within pathology and our aim is to present the possibilities of molecular detection of infectious organisms for pathological diagnosis.
- MeSH
- Molecular Diagnostic Techniques * classification methods MeSH
- Formaldehyde MeSH
- In Situ Hybridization methods MeSH
- Communicable Diseases * diagnosis etiology pathology MeSH
- Humans MeSH
- Microbiota genetics MeSH
- Nucleic Acids analysis isolation & purification MeSH
- Polymerase Chain Reaction classification methods MeSH
- High-Throughput Nucleotide Sequencing methods MeSH
- Check Tag
- Humans MeSH
- Publication type
- Review MeSH
- Keywords
- molekulární psychiatrie,
- MeSH
- Molecular Diagnostic Techniques methods MeSH
- Wastewater-Based Epidemiological Monitoring MeSH
- Congresses as Topic MeSH
- Humans MeSH
- Microbiota genetics MeSH
- Molecular Biology * trends MeSH
- Respiratory Tract Diseases epidemiology prevention & control MeSH
- Nucleic Acids analysis MeSH
- RNA Viruses pathogenicity MeSH
- High-Throughput Nucleotide Sequencing methods instrumentation MeSH
- Check Tag
- Humans MeSH
- MeSH
- Molecular Diagnostic Techniques MeSH
- Congresses as Topic MeSH
- Humans MeSH
- Molecular Biology MeSH
- Nucleic Acids * analysis MeSH
- Check Tag
- Humans MeSH
- Publication type
- News MeSH
The global risk of viral disease outbreaks emphasizes the need for rapid, accurate, and sensitive detection techniques to speed up diagnostics allowing early intervention. An emerging field of microfluidics also known as the lab-on-a-chip (LOC) or micro total analysis system includes a wide range of diagnostic devices. This review briefly covers both conventional and microfluidics-based techniques for rapid viral detection. We first describe conventional detection methods such as cell culturing, immunofluorescence or enzyme-linked immunosorbent assay (ELISA), or reverse transcription polymerase chain reaction (RT-PCR). These methods often have limited speed, sensitivity, or specificity and are performed with typically bulky equipment. Here, we discuss some of the LOC technologies that can overcome these demerits, highlighting the latest advances in LOC devices for viral disease diagnosis. We also discuss the fabrication of LOC systems to produce devices for performing either individual steps or virus detection in samples with the sample to answer method. The complete system consists of sample preparation, and ELISA and RT-PCR for viral-antibody and nucleic acid detection, respectively. Finally, we formulate our opinions on these areas for the future development of LOC systems for viral diagnostics.
- MeSH
- Biosensing Techniques MeSH
- Equipment Design MeSH
- DNA, Viral analysis MeSH
- Enzyme-Linked Immunosorbent Assay MeSH
- Real-Time Polymerase Chain Reaction MeSH
- Lab-On-A-Chip Devices * MeSH
- Humans MeSH
- Microfluidic Analytical Techniques instrumentation MeSH
- Nucleic Acids analysis MeSH
- Virus Diseases diagnosis MeSH
- Point-of-Care Systems MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
- Review MeSH
BACKGROUND AND OBJECTIVES: Utilisation of the one-step nucleic acid amplification (OSNA) molecular biology method for the detection of the metastatic involvement of sentinel lymph nodes (SLNs) in endometrial cancer (EC) patients. A comparison with histopathological ultrastaging and a description of the clinical consequences. METHODS: Surgically treated EC patients underwent detection of SLNs. Nodes greater than 5 mm were cut into sections 2-mm thick parallel to the short axis of the node. Odd sections were examined according to the OSNA method, while even ones according to an appropriate ultrastaging protocol. Nodes less than or equal to 5 mm were cut into halves along the longitudinal axis with one half examined according to the OSNA method and the other half by ultrastaging. RESULTS: Fifty-eight patients were included and 135 SLNs were acquired. Both ultrastaging and OSNA agreed on 116 results. According to the OSNA method, 20.69% more patients were classified into International Federation of Gynecology and Obstetrics (FIGO) stage III. When comparing the results of the OSNA method to the conclusions of ultrastaging as a reference method, sensitivity of 90.9%, specificity of 85.5% and concordance of 85.9% were attained. CONCLUSIONS: The results of the OSNA method showed a higher frequency of detection of micrometastases and included 20.69% more patients into FIGO stage III.
- MeSH
- Adenocarcinoma, Clear Cell genetics secondary surgery MeSH
- Sentinel Lymph Node Biopsy MeSH
- Adult MeSH
- Keratin-19 genetics MeSH
- Middle Aged MeSH
- Humans MeSH
- Lymphatic Metastasis MeSH
- Neoplasm Micrometastasis MeSH
- Survival Rate MeSH
- Biomarkers, Tumor genetics MeSH
- Endometrial Neoplasms genetics pathology surgery MeSH
- Follow-Up Studies MeSH
- Nucleic Acids analysis genetics MeSH
- Intraoperative Period MeSH
- Prognosis MeSH
- Aged, 80 and over MeSH
- Aged MeSH
- Sentinel Lymph Node pathology surgery MeSH
- Cystadenocarcinoma, Serous genetics secondary surgery MeSH
- Nucleic Acid Amplification Techniques methods MeSH
- Check Tag
- Adult MeSH
- Middle Aged MeSH
- Humans MeSH
- Aged, 80 and over MeSH
- Aged MeSH
- Female MeSH
- Publication type
- Journal Article MeSH
- Comparative Study MeSH
Surface plasmon resonance microscopy and imaging are optical methods that enable observation and quantification of interactions of nano- and microscale objects near a metal surface in a temporally and spatially resolved manner. This review describes the principles of surface plasmon resonance microscopy and imaging and discusses recent advances in these methods, in particular, in optical platforms and functional coatings. In addition, the biological applications of these methods are reviewed. These include the detection of a broad variety of analytes (nucleic acids, proteins, bacteria), the investigation of biological systems (bacteria and cells), and biomolecular interactions (drug-receptor, protein-protein, protein-DNA, protein-cell).
- MeSH
- Bacteria isolation & purification ultrastructure MeSH
- Equipment Design MeSH
- Humans MeSH
- Protein Interaction Mapping instrumentation methods MeSH
- Microscopy instrumentation methods MeSH
- Nucleic Acids analysis MeSH
- Surface Plasmon Resonance instrumentation methods MeSH
- Proteins analysis MeSH
- Animals MeSH
- Check Tag
- Humans MeSH
- Animals MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
- Review MeSH
This review is focused on the reaction of 1,2-diols with ligand complexes of six-valent osmium [Os(VI)L] (where L is a nitrogenous ligand) and possibilities of electrochemical analysis of yielded products. A number of biologically important molecules, such as mono-, oligo- and polysaccharides, RNA and glycoproteins, belong to compounds containing 1,2-diol in their structure. These compounds react with Os(VI)L yielding relatively stable ligand osmate esters which are electrochemically active and suitable to the electrochemical analysis. The ligand osmate esters give redox peaks at the mercury and carbon electrodes. The redox peaks are due to the electrochemical reduction or oxidation of osmium atoms. The osmate esters with some ligands give catalytic peaks at the mercury electrodes. The catalytic peak is due to the catalytic hydrogen evolution and is very sensitive. With the catalytic peak it is possible to measure picomolar concentrations in some cases. We have used reactions of Os(VI)L for the analysis of glycans and glycoproteins in relation to their great importance in biomedicine.
- Keywords
- biodozimetrie,
- MeSH
- Electrochemical Techniques methods MeSH
- Hematopoiesis radiation effects MeSH
- Histones analysis radiation effects MeSH
- Immunophenotyping methods MeSH
- Radiation, Ionizing * MeSH
- Humans MeSH
- Micronucleus Tests methods MeSH
- Nucleic Acids analysis MeSH
- Oxidative Stress radiation effects MeSH
- DNA Damage * radiation effects MeSH
- Flow Cytometry methods MeSH
- Radiation Exposure * analysis MeSH
- Check Tag
- Humans MeSH
- Publication type
- Research Support, Non-U.S. Gov't MeSH
- Comparative Study MeSH
Sample preparation plays an important role in the DNA analysis workflow. Real samples often include a complex matrix, such as blood and other bodily fluids, or exogenous impurities, e.g., from the scene of crime. Most of the common nucleic acids isolation techniques are based on extractions; however, isotachophoretic focusing has recently attracted some interest for its simplicity and potential for very high enrichment factors and ease of automation. Here, we report on the use of a commercial isotachophoretic instrument for optimization of DNA focusing and preparative fraction collection. In order to achieve a high recovery and enrichment, experimental factors including electric current, sample amount and matrix were investigated experimentally as well as by computer simulation. The sample of a DNA ladder was injected in 30 μl volume and after ITP focusing the DNA zone was recovered using an on-column micropreparative collection valve. The DNA content in the collected sample was verified by fluorescence spectrometry and chip capillary electrophoresis with fluorescence detection.
