Super-resolution microscopy is frequently employed in the life sciences, but the number of 18 freely accessible and affordable microscopy frameworks, especially for single particle tracking 19 photo-activation localization microscopy (sptPALM), remains limited. To this end, we designed 20 the miCube: a versatile super-resolution capable fluorescence microscope, which combines high 21 spatiotemporal resolution, good adaptability, low price, and easy installation. By providing all 22 details, we hope to enable interested researchers to build an identical or derivative instrument. 23The capabilities of the miCube are assessed with a novel sptPALM assay relying on the 24 heterogeneous expression of target genes. Here, we elucidate mechanistic details of 25 catalytically inactive Cas9 (dead Cas9) in live Lactococcus lactis. We show that, lacking specific 26 DNA target sites, the binding and unbinding of dCas9 to DNA can be described using simplified 27 rate constants of kbound free = 30-80 s -1 and kfree bound = 15-40 s -1 . Moreover, after providing 28 specific DNA target sites via DNA plasmids, the plasmid-bound dCas9 population size decreases 29 with increasing dCas9 copy number via a mono-exponential decay, indicative of simple 30 disassociation kinetics. 31 1.31 ± 0.05 µm 2 /s). This value is consistent with simulated cytoplasmic diffusion of dCas9-PAmCherry2