5'-Deoxythymidine is a synthetic nucleoside analog of thymidine, featuring a deoxyribose sugar modified by the absence of the 5'-hydroxyl group. This structural modification prevents its phosphorylation by cellular kinases, making it unsuitable as a substrate for DNA synthesis. It is primarily employed in biochemical and pharmacological research to investigate nucleoside transport across cell membranes, as it is recognized by nucleoside transporters (e.g., ENT1) but cannot be metabolized further. Additionally, it serves as a tool in studies of thymidine kinase activity, nucleoside metabolism, and as an internal standard in analytical assays for nucleosides. Its use extends to the synthesis of modified oligonucleotides or probes for exploring DNA-protein interactions and enzyme inhibition mechanisms without chain elongation.