Using microradian X-ray diffraction, we investigated the crystal structure of convectively assembled colloidal photonic crystals over macroscopic (0.5 mm) distances. Through adaptation of Wilson’s theory for X-ray diffraction, we show that certain types of line defects that are often observed in scanning electron microscopy images of the surface of these crystals are actually planar defects at 70.5 angles with the substrate. The defects consist of two parallel hexagonal close-packed planes in otherwise face-centered cubic crystals. Our measurements indicate that these stacking faults cause at least 10% of stacking disorder, which has to be reduced to fabricate high-quality colloidal photonic crystals.