ID8 / ID8-luc Orthotopic Mouse Model Service for Ovarian Cancer

The ID8 / ID8-luc orthotopic mouse model of ovarian cancer engrafts spontaneously transformed murine ovarian surface epithelial cells into immunocompetent C57BL/6 mice, where slow-burn peritoneal carcinomatosis, omental colonization, and hemorrhagic ascites develop over six to ten weeks under a fully intact immune system and are tracked quantitatively by luciferase bioluminescence. Alfa Cytology delivers this platform as an integrated preclinical service: ID8 / ID8-luc cell stewardship under the line’s classical low-serum culture conditions, ovarian bursa or intraperitoneal engraftment, dose-calibrated imaging schedules with optional ascites management, checkpoint and TGF-β/PD-L1 combination dosing, and endpoints spanning tumor nodules, ascites, immune profiling, and survival analysis.

Overview of ID8 / ID8-luc Orthotopic Mouse Model for Ovarian Cancer

ID8 is the most widely used syngeneic mouse model of ovarian cancer, created by Katherine Roby and colleagues in 2000 through the spontaneous malignant transformation of mouse ovarian surface epithelial cells in continuous culture — the first practical route to transplantable murine ovarian tumors in immunocompetent C57BL/6 hosts. Its biology is honest about its limits: ID8 does not recapitulate the TP53 mutation or the genomic instability that define human high-grade serous carcinoma, and critics have rightly noted the absence of these hallmarks. Yet what ID8 does reproduce is the disease’s geography. After intraperitoneal injection, cells attach to the peritoneal surface — vascular and avascular alike — within 48 hours; new vessels appear between weeks 4 and 8; hemorrhagic ascites begin to accumulate from week 8; and leukocyte infiltration, dominated by T cells and macrophages, is already measurable at week 2 before waning as tumors mature. The omentum becomes the primary site of dissemination, accumulating the bulk of tumor mass exactly as in patients, and the slow tempo — original clonal lines averaged 114 days to death at 5×10⁶ cells — gives the immune system time to engage, making ID8 the workhorse for studying tumor-immune interactions over realistic timescales.

Fig 1. Fluorescence imaging of orthotopic and metastatic ID8-Defb29-VEGF tumors showing white-light view with primary tumor (1°, red arrows) and metastatic deposits, 800 nm DPA-713-IRDye800CW signal in the primary tumor, 700 nm autofluorescence, and merged channels localizing kidney (K), alongside subcutaneous ID8 reference tumors—demonstrating macroscopic surgical orthotopic implantation of ID8 tumor fragments onto the ovary for metastatic ovarian cancer imaging studies. (Foss, Catherine A., et al., 2024)

Luciferase tagging transformed the model’s logistics. ID8-luc cells, generated by retroviral pFB-neo-luciferase transduction with G418 selection, establish peritoneal tumors in roughly 83% of C57BL/6 mice by three to four weeks at the standard 5×10⁶ cell challenge, with near-universal take in NSG hosts. A landmark characterization mapped the model’s dose-behavior relationship precisely: 5×10⁶ ID8-fLuc cells produce ascites around day 65 with a median survival of 63.5 days that repeated ascites drainage extends to 73 days; 10×10⁶ cells accelerate ascites to week 8; 15×10⁶ to week 6. Luciferase transduction itself modestly accelerates in vivo growth relative to parental ID8 — a documented quirk that argues for fixed-lot usage across a study. The orthotopic variant is equally validated: 1×10⁶ cells under the ovarian bursa yield primary tumors, carcinomatosis, and ascites with mice becoming moribund around day 90, while tumor cells recovered from ascites re-injected into new hosts shorten the course to roughly 60 days — the founding logic of the in vivo-selected ID8-T6 invasive subline. Around the parental line, the community has built a variant ecosystem — ID8-Vegf-A for angiogenesis programs, ID8-ova for antigen-specific immunology, and CRISPR derivatives such as ID8-DKO (Trp53/Brca2-null) whose PD-L1 is inducible by TGF-β and IFN-γ and which underpinned bintrafusp alfa combination studies — making the ID8-luc orthotopic platform the Swiss-army system of ovarian immuno-oncology.

Cell Line Information: ID8 / ID8-luc

ID8 is a spontaneously transformed murine ovarian surface epithelial line with a documented dissemination biology, and its luciferase derivative has been quantitatively characterized in immunocompetent hosts. Core attributes follow.

Our Services

ID8’s slow tempo is its scientific strength and its operational test: with courses running six to ten weeks, small early inconsistencies echo all the way to the survival curve. Alfa Cytology meets that test with discipline built for long horizons — the line’s classical low-serum culture recipe is followed verbatim, fixed reporter lots are used across each study (respecting the documented growth difference of luciferase-tagged cells), and engraftment doses are chosen from the published dose-behavior map with pilot-run confirmation. Imaging runs on calibrated calendars, ascites management follows written drainage protocols where designs require it, and variant lines from the ID8 ecosystem — VEGF-overexpressing, OVA-expressing, p53/Brca2-null — are available as matched companions. Through nodule counts, ascites volumes, immune profiling, and survival analysis, Alfa Cytology runs the ID8 / ID8-luc orthotopic workflow as one continuous, accountable program.

Workflow of ID8 / ID8-luc Orthotopic Mouse Model Construction

The ID8-luc orthotopic workflow is engineered around the model’s slow, immune-rich biology: longer calendars, dose discipline drawn from published maps, and endpoints that harvest the peritoneal disease in full. The sequence below is the standard construction-to-analysis flow.

1. Line stewardship and reporter release: ID8 / ID8-luc stocks are maintained in the classical DMEM-4% FBS-ITS recipe, mycoplasma-tested, and released only after uniform in vitro luciferase glow is confirmed; a single fixed reporter lot is used across each study.
2. Inoculum preparation: Confluent cultures are trypsinized, washed, counted, and resuspended in sterile PBS at the study dose (5×10⁶ cells in 100-200 µL for standard intraperitoneal designs; 1×10⁶ cells for intrabursal placement; dose titration per the published 5/10/15×10⁶ map for accelerated timelines).
3. Recipient preparation: Female C57BL/6 mice (7-8 weeks, about 20-22 g) are acclimatized and body-weight stratified; NSG hosts are used where take-rate maximization is required; intrabursal arms proceed under anesthesia with aseptic surgical exposure of the ovarian bursa.
4. Route-specific engraftment: For intraperitoneal arms, the suspension is injected into the lower abdominal cavity; for orthotopic arms, 1×10⁶ cells in 5-10 µL are microinjected beneath the bursal membrane with the tract closed by brief pressure before repositioning the tract and closing in layers.
5. Engraftment verification and randomized allocation: Three to four weeks after injection, intraperitoneal D-luciferin (200 µL of 15 mg/mL) and IVIS imaging verify peritoneal establishment — expected in the great majority of syngeneic hosts — and cohorts are randomized by matched photon flux into treatment arms.
6. Longitudinal treatment and monitoring: On weekly in vivo imaging calendars, bioluminescence tracks peritoneal spread, omental accumulation, and ascites onset, with body weight, abdominal girth, and clinical condition scored in parallel; where protocols call for it, ascites is drained on a written schedule, which published experience shows can extend median survival meaningfully.
7. Endpoint and peritoneal analysis: At termination, peritoneal nodules are counted and weighed across omentum, mesentery, diaphragm, liver, and spleen surfaces; ascites volume is measured and cells recovered; ex vivo imaging localizes deposits; and flow cytometry profiles T cells, macrophages, and other infiltrates — connecting tumor burden with the immune dynamics that make ID8 uniquely informative.

Fig 2. ID8 / ID8-luc orthotopic mouse model construction workflow.

Case Study-ID8 / ID8-luc Orthotopic Mouse Model Development

Alfa Cytology has implemented the ID8 orthotopic platform in preclinical programs spanning checkpoint and bispecific antibodies, vaccine and cytokine combinations, anti-angiogenic agents, and oncolytic virotherapy. In these programs, dose-calibrated engraftment — drawn from the published 5/10/15×10⁶ behavior map and confirmed in pilot runs — set disease tempo to study needs, weekly in vivo imaging resolved peritoneal spread and omental accumulation alongside ascites onset, and written ascites-management protocols extended observation windows where designs required them. Terminal analyses combined peritoneal nodule counts, ascites volumes, ex vivo imaging, and immune-infiltrate flow cytometry. Variant lines from the ID8 ecosystem were deployed as matched companions where mechanism demanded them. Study-specific parameters and results are retained as sponsor-confidential deliverables, and representative outlines can be furnished upon request.

Why Choose Alfa Cytology?

ID8 is a slow model with a rich ecosystem — and it rewards CROs that respect both halves. Alfa Cytology’s service is organized around long-horizon discipline and variant fluency. Sponsors benefit from:

• The classical low-serum DMEM-4% FBS-ITS culture recipe followed verbatim, preserving the line’s biology.
• Fixed-lot reporter usage across studies, controlling the documented in vivo growth difference of luciferase-tagged cells.
• Engraftment dosing chosen from the published dose-behavior map (ascites at day 65/56/42 for 5/10/15×10⁶ cells).
• Fully immunocompetent C57BL/6 hosts with T-cell- and macrophage-rich infiltrates profiled from week 2.
• Written ascites-management protocols that published experience shows can extend median survival by about ten days.
• Access to the variant ecosystem — VEGF-overexpressing, OVA-expressing, and Trp53/Brca2-null companions.
• Terminal packages combining nodule mapping, ascites recovery, ex vivo imaging, and immune-infiltrate profiling.

Contact Us

Need an immunocompetent ovarian cancer model whose slow, immune-rich course matches your therapeutic timeline? Contact us to discuss engraftment route, dose calibration, and endpoint design for the ID8 / ID8-luc orthotopic platform. You can also reach out to Alfa Cytology for a customized proposal and quotation — our team will respond with a study design aligned to your program.

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