Compared with existing chemical and microbiological analysis, image analysis as non-destructive quality assessment is promising a cheaper, faster process and also possible to capture data in real time. This study is aimed to determine the reliability of image analysis for assessing fish freshness in room temperature for 15 hours. Whole Cyprinus carpio were used as samples for image analysis, sensory analysis, chemical analysis (pH and TVBN) and microbiological analysis (TPC). Image analysis was focused on eye and gill. The results showed that fish freshness assessment using image analysis was in line with quality changes based on sensory, chemical and microbiological analysis. Image analysis of the eyes, showed that Value of HSV analysis and L of L, *a, *b analysis were increased. Image analysis of the gills showed that Hue, Saturation and Value of HSV analysis and *a and *b of L, *a, *b analysis were decreased while L* value was increased.