1. The nitrogen (δ15N) and carbon (δ13C) isotope ratios of animal tissues can help identify the composition of diets and open up a myriad of ecological applications. However, consumers do not ingest or assimilate all components of food items, and it is not well understood how sampling different tissues of sources and consumers may affect isotopic values ascribed, and thereby how such variation affects derived ecological measures. 2. Utilising a simple prey–predator feeding relationship in insects, we examined isotopic differences in soft, exoskeleton and whole tissues using samples with and without lipid extraction. As a derived ecological measure, we calculated trophic discrimination factors, changes in δ15N or δ13C between source and consumer, for the different prey - predator tissue combinations. 3. Lipid extraction did not affect δ15N values and we found significant tissue differences in δ15N that varied between prey and predator. Lipid extraction enriched δ13C values in most instances, and it was only after extraction of lipids that we observed consistent depletion of δ13C in exoskeleton relative to soft tissues in prey and predator. 4. Isotopic differences between tissue types propagated marked variation in derived ecological parameters. Common sampling practice using whole tissue for prey and predator (whole : whole) resulted in a trophic discrimination factor of 0.48‰ for δ15N, compared with correct factors of 0.97‰ (soft : whole) and 2.18‰ (soft : soft) using prey soft tissue actually ingested by the predator. For δ13C, variation across discrimination factors was less, with whole : whole tissue of -0.14‰, whilst correct factors were -0.55‰ (soft : whole) and -0.04‰ (soft : soft). 5. Our results indicate tissue selection and preparation are important considerations for isotopic studies using arthropods. Lipid extraction is necessary to derive accurate δ13C values based on proteins, whilst consequences of tissue selection are likely context-dependent: In poorly defined systems where sources are isotopically similar or have larger variance our results indicate tissue selection within sources is important to avoid significant error, whether estimating trophic positions or dietary proportions using mixing models. In such cases we strongly recommend exclusion of source materials not assimilated in consumers.