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This study investigated the influence of dietary lipid source on the fatty acid (FA) composition of kidney (perirenal) adipose tissue in Suffolk rams. The objective was to evaluate the efficacy of replacing a palm-based protected fat (Megalac, high in palmitic acid) with a protected fish oil supplement, with or without the inclusion of Green Lip Mussel (GLM) extract, to reduce the content of less desirable saturated fatty acids (SFAs) and enhance the deposition of beneficial long-chain omega-3 polyunsaturated fatty acids (LC n-3 PUFAs; EPA and DHA). Twelve rams were randomly allocated to four dietary treatments: (1) Megalac (SFA control), (2) Megalac + GLM, (3) Fish Oil, and (4) Fish Oil + GLM. After the feeding period, kidney fat samples were analyzed for FA composition. The principal finding was that dietary lipid sources significantly altered the tissue FA profile. Rams fed fish oil-based diets exhibited a substantial reduction (P < 0.01) in the proportion of palmitic acid (C16:0) compared to those fed Megalac-based diets. However, the enrichment of kidney fat with LC n-3 PUFAs was minimal; only docosahexaenoic acid (DHA) showed a small but significant increase (P < 0.03) in fish oil groups, while eicosapentaenoic acid (EPA) remained at trace levels. The inclusion of GLM extract did not consistently enhance LC n-3 PUFA deposition. The fraction of "remaining fatty acids" was significantly higher (P < 0.05) in the fish oil groups, suggesting the accumulation of minor or unidentified FAs. In conclusion, protected fish oil supplementation effectively reduced the proportion of nutritionally unfavorable C16:0 in rams’ kidney fat but was largely ineffective in achieving substantial enrichment with EPA and DHA, highlighting the persistent barrier posed by ruminal biohydrogenation. These results indicate that future strategies to improve the FA profile of ruminant products require more effective methods to bypass rumen metabolism, potentially through combined approaches involving advanced protection technologies and specific rumen modifiers.