In all, 1.5% (w/v) agarose gel was prepared by dissolving agarose in 1× PBS. A few μl cell suspension was sandwiched between a No. 1.5 glass coverslip (VWR) and a thin slab of the agarose gel. The sample was then imaged.
The epifluorescence images were acquired by a Nikon Ti Eclipse microscope (Nikon Instruments, Inc.) using an oil immersion objective (1.46 NA, 100×), which spans an area of ~133 × 133 μm2 for DIC (no filter, autofluorescence) and fluorescence imaging (Ex 480–500 nm, Em 509–547 nm, exposure time 200 ms). The images were acquired using an EMCCD camera (Andor). They were processed using the NIS-Element AR software (Nikon Instruments, Inc.).