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The objective-type TIRFM used an inverted Nikon Eclipse Ti microscope equipped with a Nikon Apo objective (100X/1.49NA) and an electron multiplier type charge-coupled device camera (EMCCD, Andor iXonX3). The laser excitation used in this experiment was 488 nm (Melles Griot) to excite EGFP and 561 nm (Coherent Sapphire) to excite mCherry and tomato. Time-lapse images were obtained by exiting the sample by 488 nm (~7–8 mW) and 561 nm lasers (~0.3–0.4 mW) alternatively and imaged by an EMCCD. The time interval between the two lasers excitation was 1.5 s. The fluorescence of EGFP or mCherry was selectively collected with the corresponding bandpass filters (FF01–520/35-25 or FF02-641/75-25, Semrock) with the exposure time of 100 ms. The cells were imaged at room temperature (~25 °C) in the medium without phenol red (Gibco). Images analysis and single particles tracking were performed with Image J (NIH). Kymographs were generated using the plugin in Image J with the line width of 3 pixels.

The CQD submonolayer films were made by spin-coating the CQD in n-octane dispersions on a cover slide. An objective lens (Nikon CFI Plan Apochromat Lambda 100× oil, numerical aperture of 1.45) was used to collect real-space and k-space emission patterns of the CQD films. To image k-space (BFP), two coupled 200-mm focal plano-convex lenses were placed on the back optical path of the objective lens and projected the BFP image of the objective lens onto a sCMOS (scientific complementary metal-oxide semiconductor) camera (TUCSEN Dhyana 400BSI V2). A polarizer (Thorlabs CCM1-PBS251) was used to generate the polarization image of the BFP, and a band-pass filter (Semrock FF02-641/75-25) was used to filter out the excitation and noise light. See text S2 for the BFP image fitting details.