Considering the role of HOTAIR in tumorigenesis and treatment, as well as the efficiency, specificity, and irreversibility of antisense oligonucleotides binding to the target, the objective of the present study was to explore the 99mTc labeled antisense oligonucleotide probe targeting HOTAIR, evaluate its characteristics in vitro, and observe whether it can be used in the imaging of gliomas. This evidence concerns the gene HOTAIR and glioma.