Measuring Single Molecule Lifetimes


During the first few weeks of summer research, I’ve been working toward my main goal of studying the behavior of an inexpensive organic dye at the molecular level by integrating two techniques: single molecule spectroscopy (SMS) and time-correlated single photon counting (TCSPC). SMS lets us look at the electron transfer processes happening in individual molecules, which is important because in dye-sensitized solar cells (the eventual application of this research) the environment is heterogeneous, meaning that each molecule undergoes different processes at very different speeds. TCSPC measures fluorescence lifetime decays of individual molecules, and can detect processes that happen in picoseconds, while SMS alone can detect electron transfer events down to only the millisecond timescale.

I spent the first couple days here researching various organic dyes to decide which to focus on. After considering both the wavelength at which the dyes absorb best and the molar excitation coefficient, we ended up deciding to re-visit Rhodamine 560.

After the dye was selected, the next major step was to adjust the power of our laser setup so I could get a good “image” of individual dye molecules. This image is generated based on the number of emitted photons detected at each point on a tiny region of a slide coated with a low concentration solution of the dye. Finding a good power setting was a more complicated process than I expected, because there are many different settings that can be changed, which each affect the power differently. During this process, we also discovered that our laser is generating less power than it used to, which meant that I couldn’t base any of my settings off of those we had used in the past. Finally, I was able to get a clear image that showed individual dye molecules, and use SMS to confirm that they were actually single molecules.

After the images were generated, I began trying to collect fluorescence decay curves of individual dye molecules. Generating these curves for higher concentration solutions of the dye was fairly straightforward, since at higher concentrations, there is more than one molecule at every location, so more photons are emitted. However, at single molecule concentrations, very few photons are emitted, so the signal to noise ratio isn’t good enough to create a clear curve that can be fit to a mathematical function using our software. Over the past couple days, I’ve been taking a short break from this problem and returned to gathering SMS “blinking traces”, but will hopefully return to it soon!