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Tail labelled oligonucleotide probes for the detection of DNA-protein interactions
Pivoňková, Hana ; Němcová, Kateřina ; Horáková Brázdilová, Petra ; Havran, Luděk ; Macíčková-Cahová, Hana ; Hocek, Michal ; Fojta, Miroslav
DNA-protein interactions can be monitored via different ways. We introduce novel, fast and simple approaches in DNA-protein interaction detection based on electrochemical measurements of DNA alone (structure-sensitive DNA sensing), or DNA modified with osmium tetroxide bearing nitrogenous ligands, or measurements of redox-active moieties enzymatically attached to the end of a DNA substrate thus forming a labeled tail (by terminal transferase).
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Redox labelling of nucleic acids for analyzing nucleotide sequences and monitoring DNA-protein interactions
Fojta, Miroslav ; Havran, Luděk ; Horáková Brázdilová, Petra ; Pivoňková, Hana ; Kostečka, Pavel ; Macíčková-Cahová, Hana ; Raindlová, Veronika ; Vrábel, Milan ; Hocek, Michal
Nucleobase labelling of DNA for electrochemical sensing was attained through chemical modification of thymine bases with osmium tetroxide in the presence of nitrogenous ligands, or via enzymatic incorporation of nucleotide conjugates with redox-active moieties using labelled deoxynucleoside triphosphates. DNA hybridization, primer extension and PCR techniques were used for sequence-specific DNA assays. Tail-labelled DNA substrates were applied to monitor DNA binding by tumour suppressor p53 protein.
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Some New Approaches to the Characterization of the Carbon Paste Electrodes
Mikysek, T. ; Švancara, I. ; Ludvík, Jiří ; Vytřas, K. ; Kalcher, K.
Uhlíkové pastové elektrody mají dvě hlavní výhody: jejich povrch lze snadno reprodukovatelně obnovovat a dále, je možné je modifikovat v objemu za účelem získání specifických senzorů. Příprava těchto elektrod však závisí jednak na použitých materiálech, tedy na uhlíkovém prášku i na olejovém médiu, jednak na jejich vzájemném poměru. To umožňuje velikou variabilitu vlastností, současně ale otázka charakterizace těchto elektrod je velmi významná. Bylo zjištěno, že graf závislosti odporu elektrody na jejím složení je ve tvaru dvou protínajících se přímek. K vysvětlení této závislosti byl navržen, testován a diskutován model vycházející z představy těsně uspořádaných koulí (známé z krystalografie). Další způsob charakterizace těchto elektrod je založen na interpretaci cyklických voltamogramů nějakého dobře reverzibilního systému. Obě tyto metody jsou kompatibilní, navíc umožňují vysvětlení dalších jevů spojených s morfologií a se stárnutím těchto elektrod.
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Electrochemical detection of trace amounts of nitrated explosives
Barek, J. ; Fischer, J. ; Vyskočil, V. ; Pecková, K. ; Navrátil, Tomáš ; Josypčuk, Bohdan
Nitrated organic compounds belong to the group of the most frequently used explosives. Detection of low levels of such compounds is very attractive in these days. Due to the easy reducibility of the nitro group, voltammetry seems to be very suitable method for these purposes. For these determinations the following electrodes were used: were used hanging mercury drop electrode, meniscus modified silver solid amalgam electrode, miniaturized paste silver amalgam electrode, paste electrode on the basis of silver amalgam powder and proper organic liquid were used.
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Voltametrické stanovení sloučenin dusíku pomocí stříbrné pevné amalgámové elektrody
Daňhel, A. ; Barek, J. ; Josypčuk, Bohdan ; Pecková, K.
Silver amalgam is a good electrode material for construction of new types of electrodes. In this work a silver solid amalgam paste electrode was prepared. The pastes were mixed from a silver solid amalgam powder and a mineral oil, silicone oil, paraffin oil and tricresylphosphate, stuffed into a Teflon body of electrode. The silver paste amalgam electrodes are expected to provide new possibilities for determinations of electrochemically reducible compounds by modern voltammetric methods. The main expected advantage is the possibility of easily renewable surface as in the case of carbon paste electrodes.
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The use of prototype of multichannel device for electrochemical detection of DNA hybridization
Brázdilová, P. ; Navrátil, Tomáš ; Josypčuk, Bohdan ; Fojta, Miroslav
The article presents newly constructed 8-channel device, which enables sequential analysis realized on up to 8 different or equal sensors. The device is computer controlled. The screen-printed carbon electrodes (SPCE) were connected to the device. DNA hybridization was detected. DNA present in the sample spontaneously adsorbed on the surface of SPCE and free surface was subsequently blocked using beef serum albumin. The adsorbed DNA was then hybridized with the probe marked with biotin and streptavidin conjugated with enzyme alkaline phosphatase (SALP). This enzyme catalyzes the transformation of electrochemically inactive 1-naphtylphosphateone 1-naphtol, which anodic signal was detected DC voltammetry (DCV).
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