Digital colloid-enhanced Raman spectroscopy by single-molecule counting

Quantitative detection of various molecules at very low concentrations in complex mixtures has been the main objective in many fields of science and engineering, from the detection of cancer-causing mutagens and early disease markers to environmental pollutants and bioterror agents^1-5. Moreover, technologies that can detect these analytes without external labels or modifications are extremely valuable and often preferred⁶. In this regard, surface-enhanced Raman spectroscopy can detect molecular species in complex mixtures on the basis only of their intrinsic and unique vibrational signatures⁷. However, the development of surface-enhanced Raman spectroscopy for this purpose has been challenging so far because of uncontrollable signal heterogeneity and poor reproducibility at low analyte concentrations⁸. Here, as a proof of concept, we show that, using digital (nano)colloid-enhanced Raman spectroscopy, reproducible quantification of a broad range of target molecules at very low concentrations can be routinely achieved with single-molecule counting, limited only by the Poisson noise of the measurement process. As metallic colloidal nanoparticles that enhance these vibrational signatures, including hydroxylamine-reduced-silver colloids, can be fabricated at large scale under routine conditions, we anticipate that digital (nano)colloid-enhanced Raman spectroscopy will become the technology of choice for the reliable and ultrasensitive detection of various analytes, including those of great importance for human health.