Water immersion objective

Both the Fluorescence Recovery After Photobleaching (FRAP) and Raman measurements were performed on a Horiba LabRam HR Evolution with a 488nm Ar ion Melles Griot laser (Figure S1a). The power used for Raman measurements and the photobleaching step in FRAP was 25 mW at the sample (laser threshold tests were performed for EG to ensure this laser power did not alter the material), and the fluorescence was measured at 25 μW laser power, using an Olympus 40x water immersion objective (NA 0.8). Using the DuoScan mode (galvo-mirror positioned above the microscope objective) on the Horiba LabRam HR Evolution, the laser spot was rastered over a circular region with a diameter of 10 μm and binning on the Synapse BIUV CCD (2048 × 512 pixel) was increased to 7 for the FRAP measurements. The FRAP measurements were recorded with 1 second intervals and the photobleach step was performed for 2–3 seconds. Fluorescence was measured for ~ 100 seconds before and after the photobleach step during the FRAP measurement. The only modification required to add FRAP capabilities onto the LabRam was an external neutral density filter that can be inserted and removed from the 488 nm beam path without interruption to the LabSpec software (Figure S1b).