An image sensor is disclosed. The image sensor may include a semiconductor substrate including a first pixel group region and a second pixel group region, the first pixel group region including first pixel regions to sense a first light, the second pixel group region including second pixel regions to sense a second light, each of the first and second pixel regions arranged in n columns and m rows, a pixel isolation structure disposed in the semiconductor substrate to separate the first and second pixel regions from each other, first and second photoelectric conversion regions disposed in each of the first and second pixel regions of the semiconductor substrate, and a first separation structure disposed in each of the first pixel regions and in the semiconductor substrate between the first and second photoelectric conversion regions. The first separation structure may be spaced apart from the pixel isolation structure.