Can LUCS® 13 be used to stain chromosomes in living cells?

Yes, it can. Here is the step-by-step protocol we used to stain chromosomes with LUCS^® 13.

1. MDCK and NRK-52E kidney epithelial cell cultures were used for staining. The cultures were grown in DMEM/F12 (1:1) medium with 5% fetal bovine serum (FBS) and 2 mM glutamine;

2. The cells were seeded in 35 mm glass-bottomed dishes with a glass insert diameter of 14 mm at a rate of 2×10⁵ cells per dish;

3. One day after seeding, the cells reached ~60% confluence and were used for staining with LUCS^® 13;

4. A working 2.5 μM solution of LUCS^® 13 (x4010) was prepared in DMEM/F12 medium without sodium bicarbonate. The resulting solution was heated in a water bath to 37 °C;

5. The cultures in the dishes were washed twice with 1 ml of warm DMEM/F12 medium without sodium bicarbonate, after which they were incubated in 1 ml of 2.5 μM LUCS^® 13 solution in a thermostat at 37 °C for 30 minutes;

6. After the end of the incubation, the cells were washed twice with 1 mL of warm DMEM/F12 medium without sodium bicarbonate;

7. The stained cells were placed in 1 mL of warm DMEM/F12 medium without sodium bicarbonate for visualization on an inverted confocal microscope with a super-resolution module LSM900 (Zeiss, Germany);

8. The cells were imaged using a 63×/1.25 Plan-Neofluar objective, excitation wavelength 488 nm, and emission detection range 495-700 nm; the image was taken using the Airyscan module. During the image acquisition, the cells were thermostated at 37 °C. The staining results are shown in the images.