An analysis of simulated post- and antemortem (p.m. and a.m.) victim radiographs was performed. Existing pairs of bitewing radiographs randomly sampled from a large population of adolescents were used. Two categories of individuals were included based on dental therapy conditions (none or few simple fillings). Subtraction images were performed of pairs of bitewings originating from the same individual (identical images) and different individuals (non-identical images) within each group. Four observers were asked to match the bitewings and to assess the subtraction images with respect to identity based on the score system: 1 = ‘eliminated’, 2 = ‘possible’, 3 = ‘likely’, or 4 = ‘certain’. Three observers each mismatched the film radiographs among two out of 12 individuals without fillings. True positive identification (identified by scores 2 + 3 + 4) of subtraction images were made in 10–12 individuals out of a possible 12. The range of false positive (FP) and false negative (FN) scorings was 3–15 and 0–2, respectively. All observers were capable of correctly matching bitewings originating in the 12 individuals with amalgam fillings. By use of subtraction radiography, 12–14 identical images out of a possible 15 were assessed correctly. The range of FP and FN scorings of subtraction images in this group was 1–4 and 1, respectively. Defining only score 4 as positive identification, the sensitivity of the subtraction technique decreased from ~ 0.90 to 0.70 in both categories of individuals, while specificity increased to almost 1.00 from 0.82 vs. 0.96 in the categories with and without dental restorations. Bitewing radiographs from single individuals within a group of individuals were sufficiently identical to allow for valid identification by a strict criterion. The subtraction technique may add to the subjective matching of radiographs as a screening test in victim identification.