The energy-dispersive X-ray fluorescence spectrometer for analysis of conventional and micro-samples using pinhole collimators of various sizes is developed. The measurements can be performed in the air or, in order to decrease the absorption of long-wavelength radiation of low-Z elements, in helium atmosphere. The sample is excited by the air-cooled Rh target X-ray tube of ca. 100 μm nominal focal spot size and maximum power 75 W. The X-ray spectra of the samples are collected by thermoelectrically cooled Si-PIN detector. The tungsten pinhole collimators of the size holes from 50 to 2000 μm are placed between primary filter and analyzed sample to reduce size of analyzed area. The sample can be moved using the X– Y stage. The position of the sample is monitored by CCD camera and two laser pointers. The beam spot sizes for various collimators are evaluated by the thin-wire and knife-edge methods. Beside the beam spot sizes, the loss of radiation intensity and the changes of spectral distribution of the incident radiation caused by applying various collimators are also investigated. The sample-surface-down geometry in the designed spectrometer allows for a simple analysis of various samples: solutions, loose powders, solid samples of conventional size and micro-samples.