Resistant plant cultivars which used in breeding programs are considered one of the modern integrated management programs to reduce the usage of synthetic insecticides and environmental contamination, the present study aimed to characterize the resistant and susceptible tomato cultivars to Tuta absoluta based on biochemical and molecular levels, in Egypt. The biochemical characters of the tested tomato cultivars (tomato- 86, tomato- Alissa, tomato- Fayarouz, tomato- Omniya, tomato- 036, tomato- GS) were determined colorimetrically and characterized by using native- polyacrylamide gel electrophoresis (PAGE) and agarose gel. Our results showed that there were variations highly significant in all biochemical constituents of the resistant tomato cultivar (tomato- 86) compared with the susceptible one (tomato- GS). Also, native-(PAGE) for peroxidase (POD) isoenzymes techniques of the tested tomato cultivars showed variations in protein band numbers and densities in tomato-86 resistant compared with tomato-GS susceptible to Tuta absoluta infestation. The correlation coefficient between total phenols and peroxidases in infested tomato leaves and percentages of damaged leaves with the tested insect pest was negative and highly significant, while in case of total proteins and reducing sugars in infested tomato leaves as well as lycopene contents in infested tomato fruits was positive, highly significant and significant, respectively. The correlation coefficient between tomato yield means and the infested fruit percentage with T. absoluta larvae was negative and highly significant. Respecting molecular diagnosis random amplified polymorphism DNA- polymerase chain reaction (RAPD- PCR), the results demonstrated that the presence of polymorphism in the resistant tomato cultivar (tomato- 86) compared with (tomato- GS), the most susceptible to the tested insect pest infestation