Plenoptic cameras can capture 3D information in one exposure without the need for structured illumination, allowing grey scale depth maps of the captured image to be created. The Lytro, a consumer grade plenoptic camera, provides a cost effective method of measuring depth of multiple objects under controlled lightning conditions. In this research, camera control variables, environmental sensitivity, image distortion characteristics, and the effective working range of two Lytro first generation cameras were evaluated. In addition, a calibration process has been created, for the Lytro cameras, to deliver three dimensional output depth maps represented in SI units (metre). The novel results show depth accuracy and repeatability of +10.0 mm to -20.0 mm, and 0.5 mm respectively. For the lateral X and Y coordinates, the accuracy was +1.56 μm to −2.59 μm and the repeatability was 0.25 μm.