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Utilization of micropreparative isoelectric focusing in the analysis of biological samples
Vlčková, Zuzana ; Flodrová, Dana (referee) ; Šalplachta, Jiří (advisor)
The aim of this Master Thesis is the optimization and verification of conditions of separation of proteins from biological materials. For the separation a developed technique called micropreparative isoelectric focusing is being used. The theoretical part describes analytical methods of biological materials with a focus on proteins which were used for the optimization of the developing system in general. This part of the Master Thesis also presents this method itself. Furthermore, basic characteristics of other techniques used to control the efficiency of the micropreparative isoelectric focusing are introduced. The experimental part depicts the individual steps of optimization of the micropreparative isoelectric focusing. The ideal separation procedure was firstly found out on the individual standard proteins, secondly on their mixture and lastly the optimised procedure was applied to a real biological sample. The efficiency of the micropreparative isoelectric focusing was verified by SDS-PAGE and MALDI-TOF mass spectrometry.
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Analysis of selected substances contained in the extract from elderberry twigs
Škeřík, Jan ; Flodrová, Dana (referee) ; Šalplachta, Jiří (advisor)
My diploma thesis deals with the optimization and verification of conditions for separation of proteins, which are contained in the twigs of elder (Sambucus nigra L.) using the technique RP-HPLC. The measurements were made with the use of a HPLC system with a UV-VIS detector. Column Zorbax 300SB-C18 300 4,6 x 250 mm and particles the size of 5 microns were used. The theoretical part describes the attributes and usage of elder and especially its proteins. The basic characteristic of the used HPLC technique is introduced. The possibilities of how to identify proteins are described. The practical part demonstrates the individual steps of optimization of the HPLC method applied to the mixture of standard proteins. The application of this method in the real sample is also reported. The final part of the diploma thesis is focused on the comparison of analysed samples taken in different seasons.
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Development of divergent-flow isoelectric focusing chip for separation of biological agents
Duša, Filip ; Šalplachta, Jiří ; Horká, Marie ; Lunerová, K. ; Rosenbergová, K. ; Kubíček, O.
Isoelectric focusing (IEF) is an efficient method for separation of different bacterium species. Divergent flow format of IEF (DF-IEF) enables continual separation of large volumes of samples and fractionation of the separated analytes for later analysis. The mentioned properties enabled development of DF-IEF chip for separation of H2O2 inactivated biological agents (B-agents). The chip was designed to perform separation in vertical alignment which solved issues connected with sedimentation of bacterial cells. The separation method was optimized using colored markers of isoelectric point and four different b-agents were separated using the DF-IEF chip. The obtained fractions were subsequently analyzed with matrix assisted laser desorption/ionization mass spectrometry (MALDI-MS). The mass spectra proved that the developed DF-IEF chip is an effective tool for separation of inactivated B-agents.
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Reduction Of Metal Artefacts In Ct Data With Submicron Resolution Using Dual-Target Ct
Víteček, Jakub ; Šalplachta, Jiří
The article deals with the possibility of the metal artefact reduction in computed tomography (CT) data with submicron resolution using dual-target CT. The sample is scanned twice at different acquisition parameters, at two different energy spectra. Dual-energy data are then used for easier localisation and segmentation of metal areas and the final combination of low and high-density materials. The final images are compared with the projection-based metal artefact reduction (MAR) algorithm and the commercial program VGStudio MAX 3.1. The results show good functionality of the proposed method and potential for further development.
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Utilization of micropreparative isoelectric focusing in the analysis of biological samples
Vlčková, Zuzana ; Flodrová, Dana (referee) ; Šalplachta, Jiří (advisor)
The aim of this Master Thesis is the optimization and verification of conditions of separation of proteins from biological materials. For the separation a developed technique called micropreparative isoelectric focusing is being used. The theoretical part describes analytical methods of biological materials with a focus on proteins which were used for the optimization of the developing system in general. This part of the Master Thesis also presents this method itself. Furthermore, basic characteristics of other techniques used to control the efficiency of the micropreparative isoelectric focusing are introduced. The experimental part depicts the individual steps of optimization of the micropreparative isoelectric focusing. The ideal separation procedure was firstly found out on the individual standard proteins, secondly on their mixture and lastly the optimised procedure was applied to a real biological sample. The efficiency of the micropreparative isoelectric focusing was verified by SDS-PAGE and MALDI-TOF mass spectrometry.
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Analysis of selected substances contained in the extract from elderberry twigs
Škeřík, Jan ; Flodrová, Dana (referee) ; Šalplachta, Jiří (advisor)
My diploma thesis deals with the optimization and verification of conditions for separation of proteins, which are contained in the twigs of elder (Sambucus nigra L.) using the technique RP-HPLC. The measurements were made with the use of a HPLC system with a UV-VIS detector. Column Zorbax 300SB-C18 300 4,6 x 250 mm and particles the size of 5 microns were used. The theoretical part describes the attributes and usage of elder and especially its proteins. The basic characteristic of the used HPLC technique is introduced. The possibilities of how to identify proteins are described. The practical part demonstrates the individual steps of optimization of the HPLC method applied to the mixture of standard proteins. The application of this method in the real sample is also reported. The final part of the diploma thesis is focused on the comparison of analysed samples taken in different seasons.
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