Low expression of fluorescent molecules permits simultaneous detection and spatial resolution of individual fluorescent molecules.

Total internal reflection fluorescence microscopy restricts illumination to the plasma membrane.

Proteins expressed in Xenopus oocytes.

40% of fluorescent spots showed signal from both Ir8a-mCherry and Ir84a-EGFP. By contrast, fluorescence overlap for Ir84a and Ir25a is only 5%, consistent with the expected random colocalization.

The average distance between center positions of colocalizing spots of Ir8a and Ir84a is 54.7nm.

Low expression of fluorescent molecules permits simultaneous detection and spatial resolution of individual fluorescent molecules.

Total internal reflection fluorescence microscopy restricts illumination to the plasma membrane.

Proteins expressed in Xenopus oocytes.

40% of fluorescent spots showed signal from both Ir8a-EGFP and Ir84a-mCherry. By contrast, fluorescence overlap for Ir84a and Ir25a is only 5%, consistent with the expected random colocalization.

The average distance between center positions of colocalizing spots of Ir8a and Ir84a is 47.6nm.