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Circulating tumor DNA (ctDNA) has been increasingly proposed as a biomarker for characterizing and monitoring minimal residual disease (MRD) in various solid cancers, e.g., breast cancer and lung cancer. However, such studies for intracranial gliomas remain very few. Cerebrospinal fluid (CSF) is a promising source of ctDNA but still not enough for in-depth application. In this background, here, we reported on glioma cases: 1. Tumor in-situ fluid (TISF) is the local fluid at the tumor site after surgical resection. Through an intraoperatively implanted Ommaya reservoir, we obtained it postoperatively for ctDNA analysis. 2. TISF ctDNA could reflect the existence of MRD in glioma patients although no malignancy was observed by MRI. 3. Sequential follow-up TISF ctDNA could track the molecular evolution of MRD and identify pseudoprogression (PsP) after surgery. To the best of our knowledge, it is the first evidence of TISF-based liquid biopsy for glioma, which would intrigue scholars and start new conversations in this field.