To accurately compare the abundance of a protein spots across 2-dimensional gels, it is essential to compensate for variations from external sources, such as variation in staining time. Normalization is the process of removing such variation. In this analysis, a normalized volume was used, which is a computerized construct that creates a 3-dimensional visualization of the protein spot on the basis of the intensity of the spot. For example in panel A, a spot is represented as having a low normalized volume in a nonsurvivor gel; the same spot is demonstrated in panel B. This normalized volume can be compared between gels to determine whether there is a difference in intensity (C).