The evaluation of ERK1 dimerization models using fluorescence correlation Spectroscopy and the development of analysis algorithms for single-molecule super resolution

Author

Fang Huang

Publication Date

2-1-2012

Abstract

In this dissertation, we describe three advancements in the area of fluorescence spectroscopy and microscopy. First, we have implemented Fluorescence Correlation Spectroscopy on a home-built confocal microscope. We proposed F/N analysis which stands for the ratio between average intensity and average number of moving objects in FCS as a method to detect dimerization of fluorescently tagged proteins diffusing in three dimensions. We used the technique to evaluate the dimerization hypothesis of ERK1 protein during nucleus translocation and it shows that ERK1 nucleus translocation is dimerization independent. Second, we have developed a multi-emitter analysis algorithm for single-molecule super resolution techniques that is designed to localize multiple overlapping emitters within a small sub-region. We have demonstrated its advantage compared with single emitter fitting using both simulated data sets and cell data sets. Fitting processes are implemented in GPU using CUDA to further improve the speed of analysis. Third, we proposed a robust, consistent and reliable rejection algorithm using the log-likelihood ratio as a test statistic and have demonstrated its advantage by comparing to two commonly used test statistics in the literature. To predict the correct distribution of LLR under Poisson noise when expectation values approaches zero, we proposed a hypothesis which is used to obtain the correct distribution of LLR and together with the hypothesis we proposed a empirical function which could be used to obtain the distribution.

Degree Name

Physics

Level of Degree

Doctoral

Department Name

Physics & Astronomy

First Committee Member (Chair)

Diane, Lidke

Second Committee Member

Prasad, Sudhakar

Third Committee Member

N/A

Language

English

Keywords

Fluorescence spectroscopy, Fluorescence microscopy, Light beating spectroscopy, Computer algorithms, Dimers, Molecules, Cellular signal transduction.

Document Type

Dissertation

Recommended Citation

Huang, Fang. "The evaluation of ERK1 dimerization models using fluorescence correlation Spectroscopy and the development of analysis algorithms for single-molecule super resolution." (2012). https://digitalrepository.unm.edu/phyc_etds/27