Detection of residual tumor during resection of glial brain tumors remains a challenge because of a low inherent contrast of adjacent edematous brain, the surrounding infiltration zone, and the solid tumor. Therefore, new technologies that may facilitate an intraoperative analysis of the tissue at the resection edge are of great interest to neurosurgeons. For ex vivo imaging of gliomas in a mouse model and human biopsy specimens of brain tumors and nervous system tissue we have used a time-domain Sirius 713 Tomograph with a central wavelength of 1,310 nm and a coherence length of 15 microm equipped with a mono mode fiber and a modified optical coherence tomography (OCT) adapter containing a lens system for imaging at a working distance of 2.5 cm. A spectral-domain tomograph using 840 nm and 930 nm superluminescence diodes (SLD) with a central wavelength of 900 nm was used as a second imaging modelity. Both time-domain and spectral-domain coherence tomography delineated normal brain, the infiltration zone and solid tumor in murine intracerebral gliomas. Histological evaluation of H&E sections parallel to the optical plain demonstrated that tumor areas of less than a millimeter could be detected and that not only solid tumor, but also brain invaded by a low-density single tumor cells produced an OCT signal different from normal brain. Spectral-domain OCT (SD-OCT) demonstrated a significantly more detailed microstructure of tumor and normal brain up to a tissue depth of 1.5-2.0 mm, whereas the interpretation of time-domain OCT (TD-OCT) was difficult at a tissue depth >1.0 mm. Because of rapid scanning times SD-OCT data could be acquired as 3D data maps, which allowed a multi-planar analysis of the tumor to brain interface. Similar to our findings in experimental gliomas, images of human nervous system tissue acquired using SD-OCT showed a characteristic signal of normal brain tissue and a detailed microstructure of tumor parenchyma. Spectral-domain OCT of experimental gliomas and human brain tumor specimens differentiates solid tumor, diffusely invaded brain tissue, and adjacent normal brain based on microstructure and B-scan signal characteristics. In conjunction with the rapid image acquisition rates of SD-OCT, this technology carries the potential of a novel intraoperative imaging tool for the detection of residual tumor and guidance of neurosurgical tumor resections.