 Response & Interference
| The spectra on the left show an example
of ratiometric oxygen sensing using PEBBLE fluorescence
in solution. Oregon Green can be seen to stay constant
as the Ruthenium dye changes its intensity in response
to varying amounts of oxygen - from Nitrogen bubbled
solution on the top (no oxygen) to air saturated water
in the middle, to Oxygen saturated solution on the
bottom.
Each of these peaks separately changes in intensity
due to varying amounts of PEBBLEs present in the
solution, or due to changes in the intensity of the
excitation light. But, the ratio of the two peaks
does not change with either of these things. So,
because each PEBBLE contains both dyes, an
internal reference for changes in excitation or PEBBLE
concentration is always carried along with each of these
nano-devices. |
|
Ratiometric fluorescence spectra of Oregon Green-488
(peak ~525 nm) and Ru-DPP (peak ~610 nm) in the presence
of three levels of oxygen in aqueous solution.
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| Another serious problem of free dyes
which PEBBLEs avoid is interference by things like
non-specific protein binding (which happens a lot in a
cellular environment). This plot shows that the Ru/Oregon
Green fluorescence ratio goes up with increasing amounts
of Bovine Serum Albumin (BSA) present in the solution,
while the PEBBLE response stays constant - which is the
desired result since the amount of Oxygen in solution is
not changing. This illustrates the way in which
the PEBBLE matrix protects the dyes from their
environment. The converse is also true: The PEBBLE
matrix protects the cells from any potentially harmful
chemicals used for sensing, so a much wider range of
compounds can be used than with free dyes. |
|
Plot showing BSA interference with free dyes, but not
with PEBBLE Oxygen sensors.
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