Enrichment of immunoaffinity technique to capture Lil3 proteins

Abstract

The experiment in this thesis is set out to learn and understand the biology or chemistry basis and to apply different techniques used in protein biochemistry research as well as to observe the results in practice. Among the various methods of studying proteins, affinity method is regarded as one of the most effective means of purifying proteins as a result of its high degree of specificity. This experiment has demonstrated how immunoaffinity technique can be used to capture Lil3 proteins from a crude source of solubilized thylakoid membrane. Immobilization method by coupling Lil3 antibody directly onto Toyopearls beads support was performed and the coupling efficiency was evaluated. The Lil3 proteins captured by antibody column were eluted with denaturing and nondenaturing elution buffers. Regeneration and reuse of the immobilized antibody-coupled beads column have been conducted for a few times, in an attempt to conserve the limited supply antibody and the economic feasibility, including time and expense. In addition, a brief analysis of protein membrane complexes and antibody was also provided.