**4. Conclusion**

The imaging promoter assay of the *acaA* promoters 1 and 3 by bioluminescence microscopy and the histochemical and β-galactosidase promoter assays yielded similar results in the evaluation *acaA* promoter activity in the different stages of *Dictyostelium discoideum* development. Moreover, we found that obtaining an overall picture of promoter activity spatiotemporally during the entire developmental process is possible with the bioluminescence imaging promoter assay. However, spectral unmixing is required to effectively normalize the different promoter activities. We also found that the imaging promoter assay is not significantly affected by autofluorescence.
