Table 1. The characteristics of fluorescent proteins used in the study.
| Protein | EC (M−1cm−1) | QY | Abs (nm) | Ex (nm) | Em (nm) | Relative brightness | Förster distance from mRuby2 (nm) | Fluorescence lifetime of purified proteins (ns) | Fluorescence lifetime in cells (ns) |
|---|---|---|---|---|---|---|---|---|---|
| mRuby2 | 113,000* | 0.38* | 559* | 600* | 100 | — | 2.58 | 2.52 | |
| mCherry | 78,000 | 0.17 | 585 | 585 | 611 | 31 | — | 1.40 (79%), 2.90 (21%) | 1.54 |
| 72000† | 0.22† | — | — | — | — | — | |||
| mCherry (I202Y) | 32,000 | 0.02 | 590 | 592 | 620 | 2 | 4.9 | 0.52 | 0.45 |
| 79,000‡ | 0.03‡ | — | — | — | — | — | — | — | |
| mCherry (I202T) | 42,000 | 0.04 | 587 | 587 | 617 | 4 | 5.1 | 0.60 | 0.52 |
| 79,000‡ | 0.05‡ | — | — | — | — | — | — | — |
Extinction coefficients were measured by the alkaline denaturation method (see Methods). The quantum yields were determined using Rhodamine 101 in EtOH as a reference (see Methods). Abs: absorption maximum, Ex: excitation maximum, Em: emission maximum. *,†,‡Data from other studies13,14,20. For purified mCherry, because the single exponential does not fit the fluorescence lifetime curves, we fitted the curves to a double exponential function convolved with an instrument response function5. Since the measurement of EC and QY could be operation sensitive, we carried out the side-by-side measurement of mCherry as a control and confirmed that the values are comparable with the previous report13.