American researchers have developed a probe for marking biomolecules that begins to fluoresce only when it is “switched on” by binding. As reported in the journal Angewandte Chemie, the reaction takes place very quickly and the difference in brightness between the “on” and “off” states is two orders of magnitude bigger than for conventional activatable probes.
Marking biomolecules in living cells with fluorescent probes is a well-established technique. New research possibilities open up when these probes are combined with bioorthogonal reactions. Such reactions can occur inside a living system without disrupting normal biochemical processes.
This makes it possible to generate “turn-on” probes: a bioorthogonal reaction binding partner is bound to the biomolecule of interest (without affecting it) and acts as an anchoring site for the fluorescent probe. The probe is devised so that its fluorescence is significantly increased when it binds to the anchoring site. Because the probes not bound to the target fluoresce far less, background fluorescence is reduced. This eliminates the need for complex washing procedures that delay observation of the cells.
For all of this to work, the probe system must work without a toxic catalyst, react quickly to allow for time-resolved observation of biological processes, and fluoresce very strongly after being “turned on” to maximize the signal–strength relative to the background. It has not previously been possible to meet all of these requirements in one system.
A team led by Ralph Weissleder at Massachusetts General Hospital and Harvard University has now developed a system that fits the bill: an unusually bright, fast reacting, biocompatible probe system with a large difference between the switched on and switched off states.
The new probe consists of two components: The first is a fluorescent dye called BODIPY (boron dipyrromethene), a three-ring system with a subunit made of one boron, two nitrogen, and two fluorine atoms. The second component is a tetrazine molecule, a six-membered ring containing four nitrogen and two carbon atoms. Tetrazine quenches the fluorescence of BODIPY, which passes incoming energy off to the tetrazine component without radiation instead of fluorescing.
The reaction destroys the probe’s tetrazine group, turning off the quenching of the fluorescence and allowing the BODIPY molecule to glow an intense green. The researchers recorded fluorescence over a thousand times stronger than that of the probe in the “off” state. This is two orders of magnitude stronger than all previously described turn-on probes.
The success of this system is due to the particularly strong fluorescence quenching made possible by the special electronic constellation and spatial arrangement of the BODIPY and tetrazine components relative to each other.About the Author
Ralph Weissleder | Angewandte Chemie
Unravelling the genetics of fungal fratricide
16.10.2018 | Uppsala University
Fungal weapon turns against the maker
16.10.2018 | Leibniz-Institut für Naturstoff-Forschung und Infektionsbiologie - Hans-Knöll-Institut (HKI)
Augsburg chemists present a new technology for compressing, storing and transporting highly volatile gases in porous frameworks/New prospects for gas-powered vehicles
Storage of highly volatile gases has always been a major technological challenge, not least for use in the automotive sector, for, for example, methane or...
When we put water in a freezer, water molecules crystallize and form ice. This change from one phase of matter to another is called a phase transition. While this transition, and countless others that occur in nature, typically takes place at the same fixed conditions, such as the freezing point, one can ask how it can be influenced in a controlled way.
We are all familiar with such control of the freezing transition, as it is an essential ingredient in the art of making a sorbet or a slushy. To make a cold...
Thin organic layers provide machines and equipment with new functions. They enable, for example, tiny energy recuperators. In future, these will be installed...
Das Zusammenspiel aus Struktur und Dynamik bestimmt die Funktion von Proteinen, den molekularen Werkzeugen der Zelle. Durch Fortschritte in der...
New measurement method allows researchers to precisely follow the movement of individual molecules over long periods of time
The function of proteins – the molecular tools of the cell – is governed by the interplay of their structure and dynamics. Advances in electron microscopy have...
16.10.2018 | Event News
02.10.2018 | Event News
01.10.2018 | Event News
16.10.2018 | Life Sciences
16.10.2018 | Physics and Astronomy
16.10.2018 | Event News