T98G Orthotopic Mouse Model Service for Brain Cancer

The T98G orthotopic mouse model delivers a hyperpentaploid human glioblastoma platform characterized by PTEN and TP53 mutations, hTERT promoter activation, and a distinctive capacity for viable G₁ arrest under nutrient deprivation, enabling rigorous preclinical evaluation of chemotherapeutic and targeted agents within the immunodeficient murine brain. Alfa Cytology designs, validates, and manages this human xenograft glioma system with integrated small-animal MRI and comprehensive neuropathological endpoints, supplying researchers with reproducible preclinical data to advance drug discovery programs.

Overview of T98G Orthotopic Mouse Model for Brain Cancer

The T98G orthotopic model is established by stereotactically implanting T98G human glioblastoma cells into the striatum of immunodeficient nude mice, generating an intracranial tumor that recapitulates the dense cellularity, nuclear polymorphism, and infiltrative growth patterns characteristic of human glioblastoma multiforme. Originally isolated from a 61-year-old Caucasian male patient, the T98G line exhibits a hyperpentaploid karyotype with a modal chromosome number of 128–132, representing a polyploid variant of its parental T98 line. The model displays anchorage-independent growth in vitro while retaining the unique capacity to enter viable G₁ arrest under stationary-phase conditions, offering a distinctive substrate for studying proliferation control alongside tumor progression.

Fig 1. Procedure for establishing an in situ glioma model. (Iturrioz-Rodríguez, Nerea, et al., 2024)

Histologically, T98G-derived orthotopic tumors display features consistent with high-grade glioma, including elevated mitotic activity, dense cellularity, and florid microvascular proliferation. The cell line harbors inactivating PTEN mutations and DNA-binding domain mutations in TP53, alongside a C250T activating mutation in the hTERT promoter that drives telomerase-mediated immortality. With documented sensitivity to temozolomide and involvement of ABCB1-mediated drug efflux in chemoresistance, the T98G model occupies a unique position for evaluating DNA-damaging agents, telomerase inhibitors, PTEN-pathway restorers, and combination chemotherapy strategies in a human-relevant preclinical setting.

Cell Line Information: T98G

T98G is a human glioblastoma multiforme cell line originally derived from a 61-year-old Caucasian male patient. It represents a polyploid derivative of the parental T98 line and has been extensively characterized as a model of hyperpentaploid glioblastoma with PTEN and TP53 pathway alterations. Key attributes are summarized below:

Our Services

Alfa Cytology delivers comprehensive T98G orthotopic glioblastoma model services encompassing stereotactic surgical implantation in immunodeficient hosts, small-animal MRI longitudinal monitoring, and detailed neuropathological endpoint analysis with PTEN, p53, and Ki67 immunohistochemistry. Every study is conducted within fully accredited vivarium facilities under IACUC oversight, ensuring scientific precision, ethical compliance, and complete data traceability from model construction through final report delivery.

Workflow of T98G Orthotopic Mouse Model Construction

Construction of the T98G orthotopic glioblastoma model employs stereotactic intracranial injection to deliver human tumor cells into the striatum of immunodeficient nude mice, ensuring reproducible engraftment, minimal surgical morbidity, and consistent growth kinetics. The protocol integrates pre-operative analgesia, aseptic stereotactic surgery, controlled cell infusion, and longitudinal small-animal MRI monitoring, as detailed below:

1. Cell Preparation and Batch Validation: T98G cells are expanded under low-passage conditions (passage 3–8) in EMEM (EBSS) supplemented with 2 mM L-glutamine, 1% non-essential amino acids, 1% sodium pyruvate, and 10% FBS to preserve hyperpentaploid karyotype and tumorigenic potential. Viability is confirmed by trypan blue exclusion (>95% required), and anchorage-independent growth capacity is verified via soft agar colony formation assay to ensure batch consistency.
2. Animal Selection and Acclimation: Female athymic nude mice (Nu/Nu, 6–8 weeks old) are selected to accommodate human xenograft engraftment. A one-week acclimation period is observed under specific-pathogen-free housing, with daily health monitoring and body weight recording to establish individual baseline parameters.
3. Pre-Operative Analgesia and Anesthesia: Mice receive pre-operative analgesia comprising buprenorphine (0.1 mg/kg) and carprofen (5 mg/kg) subcutaneously. General anesthesia is induced and maintained with isoflurane (3% induction, 1.5–2% maintenance) delivered via nose cone, with continuous respiration and body temperature monitoring throughout the procedure.
4. Surgical Site Preparation: The scalp is shaved and depilatory cream is applied to remove fur, followed by sterilization with iodine solution. The mouse is secured in a stereotaxic frame with ear bars and a bite bar, and a single midline incision is made to expose the skull surface and identify the bregma landmark.
5. Craniotomy and Dural Exposure: A small burr hole is drilled at the predetermined stereotactic coordinates (anteroposterior +1 mm, mediolateral +2 mm relative to bregma) using a high-speed microdrill under constant saline cooling. The dura mater is carefully pierced to expose the underlying parenchyma without causing cortical trauma or hemorrhage.
6. Tumor Cell Injection: A 10 µL Hamilton syringe fitted with a glass micropipette is lowered through the burr hole to a depth of −3 mm relative to bregma, targeting the right striatum. T98G cells (1 × 10⁵) in 2 µL PBS are delivered using a micropump injector over 2 minutes at a constant infusion rate. A single-cell suspension is essential to prevent clumping and ensure uniform tumor seeding within the striatal parenchyma.
7. Needle Retraction and Wound Closure: The needle is left in place for an additional 2 minutes to prevent retrograde flow along the injection tract, then withdrawn slowly over 1 minute to minimize cell tracking. The scalp incision is closed with surgical sutures or tissue adhesive, and the mouse is transferred to a heated recovery chamber.
8. Post-Operative Care and Monitoring: Mice receive carprofen-supplemented drinking water (33 µg/mL) for 72 hours post-surgery and are monitored twice daily for neurological deficits, body weight, and wound integrity. Animals are allowed to recover for a minimum of 3 days before initiation of any imaging or treatment protocol.
9. MRI Monitoring and Tumor Assessment: Starting at day 7–14 post-implantation, tumor establishment and growth are monitored via small-animal MRI (T2-weighted or contrast-enhanced T1-weighted sequences). Tumor volumes are quantified by region-of-interest segmentation, and neurological function is scored weekly to generate longitudinal disease progression curves for each animal.
10. Endpoint Analysis and Neuropathology: At study termination, mice are humanely euthanized and brains are excised, weighed, and processed for formalin-fixed paraffin embedding. Coronal serial sections are stained with H&E for morphological assessment, and immunohistochemistry is performed for Ki67, PTEN, p53, CD31, and human-specific cytokeratins to confirm tumor origin, proliferative index, pathway status, vascular density, and species fidelity.

Fig 2. T98G Orthotopic Mouse Model construction workflow.

Case Study-T98G Orthotopic Mouse Model Development

In a representative preclinical engagement, the T98G orthotopic model was utilized to evaluate the efficacy of a novel telomerase inhibitor in combination with temozolomide against PTEN-deficient glioblastoma. Following stereotactic implantation and confirmation of engraftment by small-animal MRI within the first two weeks, cohorts were randomized to receive either the combination regimen, monotherapy arms, or vehicle control. Longitudinal MRI revealed distinct tumor volume trajectories across treatment groups, with the combination cohort demonstrating attenuated expansion relative to both monotherapy and control arms. Terminal neuropathological analysis showed reduced intracranial tumor burden, diminished microvascular proliferation, and decreased Ki67 proliferation indices in treated animals, alongside preserved peritumoral brain architecture in responders. These preclinical findings informed the regimen's mechanism-of-action hypothesis and supported advancement toward subsequent pharmacology and toxicology studies.

Why Choose Alfa Cytology?

Partnering with Alfa Cytology for your T98G orthotopic glioblastoma program provides access to a human-xenograft-focused, quality-driven preclinical infrastructure optimized for intracranial tumor research and DNA-damage response evaluation. Our core differentiators include:

• Extensive experience with human glioblastoma xenograft model development, with optimized stereotactic surgical protocols that achieve consistent striatal engraftment and predictable tumor kinetics in immunodeficient nude mice.
• Integrated small-animal MRI platforms enabling non-invasive, quantitative longitudinal tracking of intracranial tumor volume without introducing serial sacrifice artifacts into your dataset.
• Rigorous cell banking and authentication procedures, encompassing STR profiling, mycoplasma screening, PTEN mutation confirmation, and p53 status validation to ensure model fidelity and batch-to-batch reproducibility.
• Adaptable study architectures supporting single-agent screening, combination temozolomide regimens, telomerase inhibitor evaluation, PTEN-pathway drug profiling, and ABCB1 efflux pump inhibitor assessment tailored to your therapeutic modality.
• Comprehensive neuropathological endpoint capabilities including digital histopathology, immunohistochemistry for PTEN, p53, and proliferation markers, multiplex immunofluorescence, and human-specific marker confirmation to generate mechanistic insights alongside efficacy data.
• A dedicated scientific project management team providing transparent milestone reporting, timeline accountability, and direct consultation from protocol design through final data package delivery.

Contact Us

Whether your program targets glioblastoma with a novel temozolomide combination, a telomerase inhibitor, a PTEN-pathway restorer, or an ABCB1 efflux pump modulator, Alfa Cytology is positioned to accelerate your preclinical development with our T98G orthotopic glioblastoma expertise. Contact us today to discuss your study design, review our imaging and surgical capabilities, and receive a customized proposal aligned with your scientific objectives and timeline. Our neuro-oncology team looks forward to collaborating with you to generate the robust, translationally relevant data your pipeline demands.

For research use only. Not intended for any clinical use.