- Nanosensors probe pH in cells
By exploiting surface-enhanced Raman scattering, scientists from Harvard University Medical School in Boston and from the Institute for Molecular Biology and Bioinformatics in Berlin have shown that gold and silver nanoaggregates can probe the pH of live cells in microscopic detail. They also demonstrated that a two-photon approach can extend the technique to a wider pH range. These methods could allow a better understanding of the trafficking, recycling and recruitment of intra- and extracellular molecules, report the researchers.
Surface-enhanced Raman scattering provides information on molecular vibrational modes. In this technique, gold and silver nanoparticles generate surface fields that enhance the signal. The two-photon version — surface-enhanced hyper-Raman scattering — can rival the intensity of fluorescence. Both Raman techniques can probe the environment of the nanoparticles, including cellular molecules in the vicinity of the particles.
From earlier work, the researchers knew that coating nanoparticles with the acid pMBA results in pH-sensitive Raman spectra. In their recent work, they calibrated readings using solutions of known pH and showed that the ratio of different molecular mode spectral peaks tracked pH from 4.5 to 8.5.
To measure cellular pH on a microscopic scale, researchers used gold nanoparticles functionalized to be sensitive to pH. After the nanoparticles were ingested, the researchers mapped the pH inside cells using the Raman peaks of the nanoparticles (spectra in upper right). Reprinted with permission of the American Chemical Society.
They tested the technique by adding pMBA-coated gold nanoaggregates 80 to 150 nm in size to a cell culture and waiting 4.5 h for them to be incorporated into endosomes. They mapped the surface-enhanced Raman spectra of the cells using a custom setup consisting of an 830-nm excitation laser, taking spectra using 1-s acquisition times and 2-μm steps.
As reported in the September issue of Nano Letters, they found that the pH varied from 6.8 to 5.4 — the lower detection limit of their nanosensor. These results agreed with the acidity expected given the time that had elapsed.
They also performed two-photon probing using a 1064-nm picosecond laser. Because the technique is more sensitive to acidity changes, the researchers could measure pH ranging from 2 to 11. They probed cells using silver nanoparticles and the two-photon technique and found that, more than 10 h after uptake, the pH varied from 4.5 to 4.9. These results agreed with those from other techniques and with the expected increasing acidity. The researchers note that parameters for two-photon surface-enhanced hyper-Raman scattering must be optimized before live-cell imaging can be performed.
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