Use cases
After #4 fat-pad orthotopic take (technique on the orthotopic page), follow long-term or resect the primary at >300 mm³ and track spontaneous lung/node mets. Covers intravasation; slower than tail vein. Parental rarely seeds bone spontaneously in nude/NOD-SCID. Not a substitute for surgical training.
Catalog: MC-h133 · Product modeling page · In-vitro spheroid
RUO. Fat-pad inoculation follows the orthotopic page and IACUC. No anesthetic doses here; not a substitute for the veterinary SOP. Do not call tail-vein or LV “spontaneous metastasis.”
Use-case overview
Use this page for the full cascade; tail vein for lung colonization only; LV for bone homing. Do not rewrite inoculum here.
| Use | Fit | Primary readout | Notes |
|---|---|---|---|
| Post-resection spontaneous lung mets / cascade | Recommended | Lung BLI/HE, nodes; log bed recurrence[2][3] | Minn: resect >300 mm³. Luc strongly recommended. |
| No resection, prolonged follow-up | Suitable | Keep primary burden separate from distant foci[1][2] | The primary may cap survival and truncate the mets window. |
| Spontaneous bone mets or experimental lung colonization | Not recommended | Parental spontaneous bone in nude is weak; tail vein is not spontaneous | Bone: LV/IT or NSG, and say so. Lung colonization: tail vein. |
Does the workflow match?
Inoculum matches orthotopic; this page adds resection and long follow-up.
| Use | vs foolproof SOP | Differences vs core SOP |
|---|---|---|
| Post-resection cascade | Core + add-ons | Orthotopic skeleton + resection window. |
| Unresected follow-up | Core + add-ons | No resection day. |
| Tail vein / LV | Other route | Skips intravasation—not spontaneous. |
Host spec
Default female nude, same as the orthotopic page[1]. NSG has a broader spontaneous spectrum (including a higher chance of bone) and must be written into the protocol[4].
| Indication / context | Sex | Strain / host | Notes |
|---|---|---|---|
| This SOP default | Female | Nude | Expect lung/nodes, not bone. |
| Broader spontaneous spectrum | Female | NSG | Wright: NSG is more permissive than nude/NOD-SCID for spontaneous bone etc.; burden arrives faster, endpoints sooner[4]. |
| Male | Not the default | — | Fat-pad models are female-standard. |
In one line
Orthotopic inject (orthotopic page) → primary growth → optional resection → long-term mets follow-up → multi-organ pathology.
D−1 / D0 timeline
1. D−7 to D0
Quarantine and #4 fat-pad inoculum on the orthotopic page.
2. Primary growth
Caliper to the set volume (often >300 mm³ before resection).
3. Resection day (optional)
Gross-complete resection; sham controls.
4. Weeks to months after
BLI/breathing/nodes; multi-organ pathology at endpoint.
Protocol overview
Scenario-specific flowchart (core engraftment skeleton with host/therapy or imaging/readout changes).
- Complete orthotopic inoculation (orthotopic page; no fat-pad recap here)
- Grow primary to set window
- Optional primary resection (often >300 mm³)
- Long-term BLI / clinical follow-up
- Multi-organ harvest
- Quantify metastatic burden
Novice pack (literature cases)
Flowchart buttons open the matching table. Full pack below, in use order.
Unpack list (resection + long follow-up; inoculum on the orthotopic page)
Fat-pad surgery, Matrigel, and 26–27G technique live on the orthotopic page—this page does not repeat Cheng’s 25% matrix. Extra items here are resection and BLI.[1][3]
| Item | Per mouse / study | Notes |
|---|---|---|
| MDA-MB-231 / Luc line MC-h133 | Start from 1 vial | Inoculum matches the orthotopic page; Luc strongly recommended—validate phenotype[2] |
| Caliper | Shared | Volume ≈ L × W² / 2; resection window often >300 mm³[3] |
| Resection kit / suture / hemostasis | Per surgical n | Clean margins; sham controls; analgesia per IACUC. No doses here |
| BLI substrate (Luc) | Per imaging SOP | Jenkins: orthotopic + multi-organ mets detectable in vivo[2] |
| Long-term SPF housing | Weeks to months | Write respiratory distress, ulcer, and weight-loss endpoints |
Expansion cases: mice vs T75 (same conservative math as orthotopic)
2×10⁶/mouse + 50% dead space. Volume, matrix, and #4 fat-pad technique are on the orthotopic page—do not invent a second recipe here.
| Mice | T75 flasks |
|---|---|
| 1 | 1 |
| 2 | 2 |
| 3 | 2 |
| 4 | 3 |
| 5 | 3 |
| 6 | 4 |
| 7 | 5 |
| 8 | 5 |
| 9 | 6 |
| 10 | 6 |
Inoculum recipe (points to the orthotopic page)
This page does not rewrite the fat-pad SOP. Spontaneous mets start after Price/Jenkins orthotopic take.[1][2]
| Item | This SOP | Literature |
|---|---|---|
| Inoculum | Open the orthotopic page for #4 fat pad | Price nude orthotopic[1]; Luc in Jenkins[2] |
| Cells / matrix | Do not pick a second dose here | Lock the same lot as the orthotopic SOP |
| Resection window | Resect the primary at a set volume | Minn: resect >300 mm³, then follow lung[3] |
| No-resection arm | Leave the primary to humane endpoint | Do not confound primary burden with mets |
Resection checklist (not the fat-pad inject card)
Minn resects primaries >300 mm³ then scores lung mets. Incomplete resection will be misread as “metastasis.” No anesthetic doses here.[3]
| Check | Pass | If fail |
|---|---|---|
| Volume window | Hit the protocol volume (often >300 mm³)[3] | Too early: mets not yet launched; too late: ulcer/burden |
| Margins | Gross-complete, hemostasis, close | Residual nodule: drop from the mets primary analysis or split out |
| Controls | Sham or volume-matched unresected | No control confounds surgery with mets |
| Post-op | Analgesia per IACUC; log weight | Failure to right or respiratory distress → vet SOP |
Culture card (MDA-MB-231 / MC-h133)
Inverted microscope (schematic 10× field): confluence is the % of the growth surface covered by cells. If the monolayer is even, that matches the fraction of the field occupied. Split/harvest at the middle panel. Click the schematic to enlarge.
Culture matches the orthotopic page. Validate that Luc does not change growth/mets[2]. Schematic, not a real micrograph.
| Item | Practice |
|---|---|
| Medium | Datasheet first. Day-of harvest is on the orthotopic page. |
| Confluence | Harvest at 85–90%; do not invent a second confluence rule. |
| Labeled line | After Luc/GFP enrichment, run parental in parallel or re-validate mets[2]. |
What success looks like (expected, not a guarantee)
Troubleshooting
Incomplete resection, too-short follow-up, or calling tail-vein lung “spontaneous” will write the wrong conclusion.
| Sign | Cause | Action |
|---|---|---|
| Rapid bed recurrence | Residual primary | Exclude from the post-resection mets primary endpoint |
| No distant lesions | Short follow-up, indolent parental, or Luc false-negative | Follow to the IACUC cap and do HE; Jenkins is in-vivo sensitive but still needs validation[2] |
| Calling bone mets a spontaneous success | Wrong host/expectation | Parental spontaneous bone in nude is weak; switch to LV or NSG and say so[4] |
| Respiratory distress / ulcer | Burden or primary | Humane endpoint now—do not wait for mets |
Monitoring log fields
Log primary volume separately from distant readouts. Scheduled whole-body BLI for Luc.
Literature case comparison
This page scores the cascade; experimental lung colonization is the tail-vein page.
| Paper | Host | Design | Readout |
|---|---|---|---|
| Price 1990[1] | Nude fat pad | Parental tumorigenicity and mets spectrum | Spontaneous mets occur; log organ spectrum by line |
| Jenkins 2005[2] | Immunodeficient | Luc orthotopic + BLI | Primary and multi-site mets detectable; validate the label |
| Minn 2005[3] | Immunodeficient fat pad | Resect primary >300 mm³, select lung mets | Yielded LM populations; do not mix LM2 tail-vein SOP onto this page |
| Wright 2016[4] | Nude / NOD-SCID / NSG | Review of spontaneous vs experimental bone mets | Parental rarely seeds bone spontaneously in nude/NOD-SCID; NSG is more permissive; use LV/IT for bone |
- 01[Prerequisite] Use the orthotopic page for ethics, cells, and #4 fat-pad injection (Price; Jenkins). This page does not repeat Cheng’s 25% matrix or nipple surgery. RUO. No anesthetic doses.
- 02[Primary] Caliper volume curves. Resection arm: gross-complete at a set volume (Minn often >300 mm³), hemostasis, sham controls[3].
Minn resects primaries >300 mm³ then scores lung mets. Incomplete resection will be misread as “metastasis.” No anesthetic doses here.[3]
Check Pass If fail Volume window Hit the protocol volume (often >300 mm³)[3] Too early: mets not yet launched; too late: ulcer/burden Margins Gross-complete, hemostasis, close Residual nodule: drop from the mets primary analysis or split out Controls Sham or volume-matched unresected No control confounds surgery with mets Post-op Analgesia per IACUC; log weight Failure to right or respiratory distress → vet SOP - 03
- 04[Endpoint] Systematic harvest of bed, lung, nodes, liver, bone. Do not treat parental spontaneous bone in nude as a positive expectation[4].
- 05[Interpretation] Slower and more heterogeneous than tail vein, but includes intravasation. Do not write i.v./LV results as spontaneous.
- 06[QC] Fix resection timing and follow-up length; independent repeats. Validate Luc phenotype[2].
Minn resects primaries >300 mm³ then scores lung mets. Incomplete resection will be misread as “metastasis.” No anesthetic doses here.[3]
Check Pass If fail Volume window Hit the protocol volume (often >300 mm³)[3] Too early: mets not yet launched; too late: ulcer/burden Margins Gross-complete, hemostasis, close Residual nodule: drop from the mets primary analysis or split out Controls Sham or volume-matched unresected No control confounds surgery with mets Post-op Analgesia per IACUC; log weight Failure to right or respiratory distress → vet SOP
Reagents / materials
| Item | Role | Conc. / dose |
|---|---|---|
| MDA-MB-231 cells (MC-h133) | Orthotopic inoculum (parameters on the orthotopic page) | 同原位 SOP（约 1×10⁶ 级；保守备瓶按 2×10⁶） |
| Luc-labeled line (strongly recommended) | Whole-body mets imaging | 验证表型后使用[[2]] |
| Anesthetic / analgesic | Surgery & welfare | 按伦理批件与兽医 SOP |
| Test drug / vehicle (optional) | Treatment arm | 按药理方案 |
Readouts
Primary growth curve, distant organ burden, BLI, survival, multi-organ pathology.
References (PubMed)
- [1]PMID 2297709 — Tumorigenicity and metastasis of human breast carcinoma cell lines in nude mice. Cancer Res (1990)
- [2]PMID 15987449 — Bioluminescent human breast cancer cell lines that permit rapid and sensitive in vivo detection of mammary tumors and multiple metastases in immune deficient mice. Breast Cancer Res (2005)
- [3]PMID 16049480 — Genes that mediate breast cancer metastasis to lung. Nature (2005)
- [4]PMID 27867497 — Murine models of breast cancer bone metastasis. Bonekey Rep (2016)
Disclaimer: RUO; IACUC required. Optimize by strain and pilot. Red tags mark weak or non-metastatic parental endpoints.