| Autor: |
Liangxing Wu, Yingrui Dai, Xiaoli Jiang, Chutima Petchprayoon, Jessie E Lee, Tao Jiang, Yuling Yan, Gerard Marriott |
| Jazyk: |
angličtina |
| Rok vydání: |
2013 |
| Předmět: |
|
| Zdroj: |
PLoS ONE, Vol 8, Iss 6, p e64738 (2013) |
| Druh dokumentu: |
article |
| ISSN: |
1932-6203 |
| DOI: |
10.1371/journal.pone.0064738 |
| Popis: |
We present the design, synthesis and characterization of new functionalized fluorescent optical switches for rapid, all-visible light-mediated manipulation of fluorescence signals from labelled structures within living cells, and as probes for high-contrast optical lock-in detection (OLID) imaging microscopy. A triazole-substituted BIPS (TzBIPS) is identified from a rational synthetic design strategy that undergoes robust, rapid and reversible, visible light-driven transitions between a colorless spiro- (SP) and a far-red absorbing merocyanine (MC) state within living cells. The excited MC-state of TzBIPS may also decay to the MC-ground state emitting near infra-red fluorescence, which is used as a sensitive and quantitative read-out of the state of the optical switch in living cells. The SP to MC transition for a membrane-targeted TzBIPS probe (C₁₂-TzBIPS) is triggered at 405 nm at an energy level compatible with studies in living cells, while the action spectrum of the reverse transition (MC to SP) has a maximum at 650 nm. The SP to MC transition is complete within the 790 ns pixel dwell time of the confocal microscope, while a single cycle of optical switching between the SP and MC states in a region of interest is complete within 8 ms (125 Hz) within living cells, the fastest rate attained for any optical switch probe in a biological sample. This property can be exploited for real-time correction of background signals in living cells. A reactive form of TzBIPS is linked to secondary antibodies and used, in conjunction with an enhanced scope-based analysis of the modulated MC-fluorescence in immuno-stained cells, for high-contrast immunofluorescence microscopic analysis of the actin cytoskeleton. |
| Databáze: |
Directory of Open Access Journals |
| Externí odkaz: |
|
