Image sensors are provided including a substrate defining a plurality of pixel regions, the substrate having a first surface and a second surface opposite the first surface. The second surface of the substrate is configured to receive light incident thereon and the substrate defines a deep trench extending from the second surface of the substrate toward the first surface substrate and separating the plurality of pixel regions from each other. In each of the plurality of pixel regions of the substrate, a photoelectric conversion region is provided. A gate electrode is provided on the photoelectric conversion region and a negative fixed charge layer covering the second surface of the substrate and at least a portion of a sidewall of the deep trench is also provided. The image sensors further include a shallow device isolation layer on the first surface of the substrate. The shallow device isolation layer defines an active region in each of the pixel regions and the negative fixed charge layer contacts the shallow device isolation layer.