In English

Two-colour fluorescence-based method for detecting virus particles in solution

Frida Ulander
Göteborg : Chalmers tekniska högskola, 2016. 56 s.
[Examensarbete på avancerad nivå]

In our globalized world people travel all over the planet in work and for leisure. But with the intensified travelling comes the effect of diseases spreading fast between continents. In case of outbreaks or epidemics, impeding spreading of diseases is of great importance and there is thus a great need of fast, reliable and sensitive virus detection methods. User-friendly detection methods could be placed at for example airports to identify virus carriers. In this thesis a novel method for detecting small amounts of viruses in solution is evaluated. The assay is based on fluorescence microscopy and by having fluorescently labelled biological molecular assemblies, called vesicles, in a solution. The role of the vesicles is to act as reporters for the unlabelled virus particles. When adding a sample containing viruses, the vesicles and viruses interact specifically, bind to each other and form complexes. By recording the movement of the vesicles with a microscope the vesicles can be tracked individually by a software. Half of the vesicles are labelled so that they are being excited by a red laser and half of the vesicles so that they are excited by a blue laser. The two subpopulations are tracked separately in the microscope. By finding events where a red and a blue particle are moving correlated, the viruses can be detected since a blue and a red particle moving together indicates that they are held together by a virus. The lowest virus concentrations to be detected was 10 pM by tracking particles and detecting colocalization events. By analysing the same data with an alternative method based on overlaying images instead of single-particle tracking, the processing time was significantly reduced, still with a limit of detection of 10 pM. However, for both analysis methods, the contrast between samples with high virus concentration and samples with no viruses was only on the order of 4. This can be partly due to limitations in the data analysis and also to the vesicles interacting unspecifically with each other.

Nyckelord: colocalization, dual-colour microscopy, virus detection, single-particle

Publikationen registrerades 2016-07-06. Den ändrades senast 2016-07-06

CPL ID: 239052

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