Quantum Dot Targeting with Lipoic Acid Ligase and HaloTag for Single-Molecule Imaging on Living Cells
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- 作者：Daniel S. Liu ; William S. Phipps ; Ken H. Loh ; Mark Howarth ; Alice Y. Ting
- 刊名：ACS Nano
- 出版年：2012
- 出版时间：December 21, 2012
- 年：2012
- 卷：6
- 期：12
- 页码：11080-11087
- 全文大小：432K
- 年卷期：v.6,no.12(December 21, 2012)
- ISSN：1936-086X
文摘
We present a methodology for targeting quantum dots to specific proteins on living cells in two steps. In the first step, Escherichia coli lipoic acid ligase (LplA) site-specifically attaches 10-bromodecanoic acid onto a 13 amino acid recognition sequence that is genetically fused to a protein of interest. In the second step, quantum dots derivatized with HaloTag, a modified haloalkane dehalogenase, react with the ligated bromodecanoic acid to form a covalent adduct. We found this targeting method to be specific, fast, and fully orthogonal to a previously reported and analogous quantum dot targeting method using E. coli biotin ligase and streptavidin. We used these two methods in combination for two-color quantum dot visualization of different proteins expressed on the same cell or on neighboring cells. Both methods were also used to track single molecules of neurexin, a synaptic adhesion protein, to measure its lateral diffusion in the presence of neuroligin, its trans-synaptic adhesion partner.