This question can be answered by considering a brief metaphor. Misconceptions of nature can arise when careful measurements are not taken on single species. Imagine you have never heard of fireflies. Your colleague places millions of fireflies in large jars (Figure 1) and asks you to study the light. After observing the jars for several minutes in the dark, you note that the intensity of light doesn’t vary greatly with time and scales linearly with the number of fireflies. You conclude fireflies continuously light due to bioluminescence. Because you did not study one firefly at a time, you failed to see a firefly blinking on and off. But how could you have known by observing millions of fireflies at a time? The probability of each firefly blinking on and off at the same exact time is very low.

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Not until scientists started studying the fluorescence of single proteins did we know that protein fluorescence was not continuous. These observations were used, in part, to invent super-resolution microscopy, which garnered the 2014 Nobel Prize in Chemistry. Thus, the study of the single entity – be it a firefly, single molecule, or nanoparticle – allows us to uncover natural phenomena that get washed out in ensemble types of experiments as well as develop game-changing techniques. Even more, from an analytical standpoint, if the observations are specific, these experiments offer access to the ultimate sensitivity in analysis: a limit of detection of a single molecular species.