The advantage of handheld type surgical microscope is that the size of the probe is small and light, and that both the working distance (15–30 cm) and field of view (30 deg) can be adjusted. The shortness working distance will minimize the loss of light source energy. However, the currently developed handheld type surgical microscope is still large, heavy, and uses relatively high energy (600 mW/cm2). To address the aforementioned problems, this study aimed to develop a pen-type surgical fluorescence microscope that is compact, portable, and has an adjustable beam angle and working distance. These features enable real-time diagnosis. The pen-type probe consists of a laser diode, CMOS camera, light source brightness control device, filter, and power switch. The IR-cut filter inside the CMOS camera was removed to facilitate transmission of the fluorescence emission wavelength. In addition, a long-pass filter was attached to the camera so that the external light source was blocked and only the fluorescence emission wavelength was allowed to pass through. The performance of the pen-type probe was tested through a large animal experiment. Indocyanine green (2.5 mg/kg) was injected into a pig's vein. Fluorescence emission of 805-830 nm was achieved by irradiating an external light source (785 nm and 4 mW/cm2), and liver-uptake occurred after 2.4 min. The designed pen-type probe was capable of sufficiently fluorescence expression through low-energy irradiation, and the pen-type probe is small and light and easy to handle by hand because both the pen-based laser device and the camera device are integrated. In addition, it is easy to adjust the working distance and field of view.