In the field of semiconductor development and failure analysis, metrology of layers such as gate oxide layer is one of the important analysis due to determine semiconductor itself characteristics. The number of requirements of metrology is increasing by using both scanning and transmission electron microscopy. High accurate metrology depends on accuracy of magnification of electron microscope. We developed accurate magnification calibration for scanning transmission microscope. This method is carried out by using micro scale specimen and silicon single crystal lattice fringe images. We achieved absolute magnification error of less than 2% for all magnification. This microscope provides high accuracy metrology for semiconductor device. We describe an automatic magnification calibration function for the high magnification range required to accurately measure features from a few to tens of nm in size.