We developed a laboratory-size three-dimensional water-window x-ray microscope using condenser and objective grazing incidence Wolter type I mirrors, an electron-impact-type x-ray source, and a back-illuminated CCD. The imaging system was improved for practical applications in life science research fields. Using a new objective mirror with reduced figure errors, a resolution limit of 3.1 line pairs/μm was achieved for two-dimensional transmission images and sub-micrometer-scale three-dimensional structures were resolved. Incorporating a cryogenic stage into the x-ray microscope, we observed biological samples embedded in ice to evaluate the usefulness of observation in the water-window region and multi-energy observation was demonstrated using an x-ray source with multiple x-ray tubes.