The white adipose tissue-specific aptamer Adipo8 can specificity bind?with mature adipocytes or tissues and inhibit adipogenesis.?In this research, we explored?the effect of Adipo8 intervention on the transcriptome in the process of adipogenesis using mRNA-level sequencing,?analyzed the mechanism of?Adipo8 in?inhibiting adipogenesis. The results showed that Adipo8 can inhibit lipid formation and downregulate PPARγ and C/EBPα in differentiated 3?T3-L1 cells. Transcriptome mRNA sequencing of 3?T3-L1 cells after Adipo8 intervention?revealed that Adipo8 might inhibit the biological function of adipogenesis by downregulating Acsl1 and Plin1 to inhibit fatty acid metabolism and PPAR signaling pathways.?After that, using Spacer18 to connect the optimized and truncated Adipo8, we constructed a bivalent aptamer Adipo8cB?and compared the affinity, biological effects, and biological stability between the aptamers in differentiated and mature 3?T3-L1 cells. At the cellular level,?the affinity, biological effects, and serum stability of Adipo8cB were verified to be superior to those of Adipo8?in 3?T3-L1 cells.?We then investigated the biological properties of Adipo8cB as a lipid-inhibiting drug in?vivo, using C57BL/6J mice with diet-induced obesity. The body weight, blood sugar, lipid levels, liver function, glucose tolerance, and other related indicators in each group of mice were observed and compared after intervention with the bivalent aptamers Adipo8cB and Adipo8. Both Adipo8cB and Adipo8 effectively prevented weight gain caused by fat accumulation in mice?with diet induced obesity, while also reducing blood lipid levels, improving glucose tolerance, and protecting against liver steatosis, moreover, Adipo8cB has a better effect than Adipo8.