Use cases
Default is 231-BR LV (Yoneda 1×10⁵ or Zhou ~1.75–2×10⁵ / 100 μL) for multifocal brain mets. Carotid follows Liu PDX technique (1×10⁵ / 50 μL)—not an MDA-231 default dose. Intracranial 1–2 μL scores local growth only. Watch JoVE—not a substitute for surgical training.
Catalog: MC-h133 · Product modeling page · In-vitro spheroid
RUO. IACUC, SPF, and LV/microsurgical training required. No anesthetic doses here; not a substitute for JoVE or the veterinary SOP. No Matrigel. Do not inject without bright-red flashback. Do not use BR on the bone-mets page.
Zhou JoVE: US-guided LV for 231-BR brain mets (video) · PMID 24637963
Use-case overview
Use LV BR (default) or carotid for hematogenous brain colonization; stereotactic for local intracranial growth. Do not import the BO/bone-page SOP.
| Use | Fit | Primary readout | Notes |
|---|---|---|---|
| Experimental BCBM (hematogenous) | Recommended | MRI count/volume, neuro signs, whole-brain pathology[1][2][4] | 231-BR; 1–2×10⁵ / 100 μL; female nude. |
| Carotid colonization across BBB | Suitable | Brain take ± less systemic spill[3] | Liu is PDX/SCID 1×10⁵ / 50 μL—pilot before using 231-BR. |
| Local intracranial growth or full cascade | Not recommended | IC skips circulation; use spontaneous orthotopic mets for the cascade | IC can screen brain-microenvironment drugs—do not call it metastasis. |
Does the workflow match?
BR LV is this page’s foolproof skeleton; carotid/IC use other cards; bone is the BO page.
| Use | vs foolproof SOP | Differences vs core SOP |
|---|---|---|
| Experimental BCBM | Same core SOP | This page’s LV. |
| Carotid | Core + add-ons | 50 μL microsurgery, not 100 μL. |
| Intracranial / spontaneous / bone | Other route | Parenchymal implant, fat pad, or BO LV. |
Host spec
LV default is female nude. Yoneda 4 wk; Zhou 6–8 wk[1][2][4]. Carotid Liu uses SCID ~8 wk[3]. Males are not the default.
| Indication / context | Sex | Strain / host | Notes |
|---|---|---|---|
| This SOP default (LV BR) | Female | Nude 6–8 weeks (4 weeks also used) | 1–2×10⁵ / 100 μL. |
| Carotid PDX technique | Female | SCID ~8 weeks[[3]] | Do not treat this host/dose as the 231-BR default. |
| Male | Not the default | — | The SOPs above are female. |
In one line
IACUC → expand BR → LV (default) or carotid/IC → MRI/clinical signs → whole-brain pathology.
D−1 / D0 timeline
1. D−7 to D−3
Quarantine female nudes; write neurologic endpoints.
2. D−1
Feed at ~80% confluence[[4]].
3. D0
BR 1–2×10⁵ / 100 μL → LV (or carotid/IC per the route cards).
4. From D+21
MRI/neuro signs; T2 ~week 3.
Protocol overview
Scenario-specific flowchart (core engraftment skeleton with host/therapy or imaging/readout changes).
- IACUC + neurologic endpoints
- Expand 231-BR (log passage; T75 on expansion table)
- Single-cell suspension (no Matrigel)
- Anesthetize; LV inject BR (bright-red flashback, 100 μL)
- MRI or BLI follow-up
- Endpoint whole-brain pathology
Novice pack (literature cases)
Flowchart buttons open the matching table. Full pack below, in use order.
Unpack list (brain mets, plan per mouse)
Default is 231-BR LV—not BO from the bone page, not a fat pad. No Matrigel. Anesthetic doses come only from the IACUC SOP.[1][2]
| Item | Per mouse / study | Notes |
|---|---|---|
| MDA-231BR (or parental control) MC-h133 | Start from 1 vial | STR/mycoplasma-qualified; log BR vs parental vs BO separately[1] |
| FBS MC100 | 10% complete medium | Regular grade |
| Serum-free DMEM or ice-cold PBS | 100 μL/mouse (LV) | Zhou PLOS: 100 μL serum-free medium[4]; Yoneda: 0.1 mL PBS[1] |
| 28G insulin syringe (LV) | 1 per mouse | Yoneda 28G; US-guided method on JoVE[1][2] |
| 33G bent needle (carotid only) | Per surgical n | Liu PDX: CCA inject, 50 μL; not the MDA-231 default dose[3] |
| Microsyringe (intracranial only) | 1–2 μL | Liu: 1×10⁵ in 1–2 μL striatum; local growth, not the cascade[3] |
| MRI / optional BLI | Shared | Multifocal T2 lesions ~week 3; contrast per imaging SOP[2][4] |
Expansion cases: mice vs T75 (stocked to LV 2×10⁵)
Inoculum recipe (by route)
Default SOP is 231-BR LV. JoVE lists 1.75×10⁵; the same group’s PLOS lists 2×10⁵. Carotid/intracranial numbers are Liu PDX—not MDA-231 defaults.[1][2][3][4]
| Route | This SOP / literature | Volume and vehicle |
|---|---|---|
| LV (default) | Yoneda 1×10⁵; Zhou JoVE 1.75×10⁵; PLOS 2×10⁵[1][2][4] | 100 μL PBS or serum-free medium; no Matrigel |
| Intracarotid | Liu PDX: 1×10⁵; female SCID ~8 wk[3] | 50 μL PBS; 33G; ~2 min push. Not the 100 μL LV volume |
| Stereotactic intracranial | Liu: 1×10⁵[3] | 1–2 μL PBS; right striatum (2 mm right of bregma, 2.5 mm deep). Local growth |
| Batch multiplier (LV) | n × 1.2 | Insulin dead space is smaller than tuberculin; load one syringe per mouse |
LV checklist (231-BR, not BO)
Do not use BR on the bone page. Do not use BO here. Inject only after bright-red flashback. JoVE uses ultrasound to raise success. Hub air is only to see the pulse—never inject air.[1][2]
| Check | Pass | If fail |
|---|---|---|
| Subline | 231-BR / Br-GFP / Br-luc | Parental is less efficient with more extracranial lesions; BO belongs on the bone page[1] |
| Flashback | Slow 100 μL after bright pulsatile return | No flashback: do not inject; do not repeat blind sticks[2] |
| Guidance | Use JoVE ultrasound if available[2] | Manual success ~50%; repeat sticks raise death[2] |
| Matrix | No Matrigel | Matrix embolizes arteries |
Carotid checklist (Liu modified ICA, PDX technique)
Dose and host are PDX/SCID—not the MDA-231BR default. Caudal CCA + ECA ligatures send cells via ICA. Watch Figure 1; not a substitute for microsurgical training. No anesthetic doses here.[3]
| Check | Pass | If fail |
|---|---|---|
| Host | Liu: female SCID ~8 wk[3] | Do not default to 4-wk nude LV hosts |
| Volume | 1×10⁵ / 50 μL PBS; 33G over ~2 min[3] | Do not inject the 100 μL LV volume |
| Direction | Block caudal backflow; ECA ligature cuts face/ear seeding[3] | Face/ear tumor = ECA distribution |
| Readout meaning | Hematogenous colonization across the BBB | Still not the orthotopic intravasation cascade |
Intracranial checklist (stereotactic, local growth)
Liu: 1×10⁵ in 1–2 μL, right striatum. Reliable take—do not extrapolate homing or BBB transit.[3]
| Check | Pass | If fail |
|---|---|---|
| Volume | 1–2 μL, slow | 10 μL injures/spills |
| Coordinates | 2 mm right of bregma, 2.5 mm deep[3] | Wrong coordinates = ventricle or extra-axial |
| What it models | Growth in brain parenchyma | Use spontaneous orthotopic for cascade; LV/carotid for hematogenous spread |
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.
Zhou: harvest near 80% confluence; resuspend serum-free[4]. Schematic, not a real micrograph.
What success looks like (expected, not a guarantee)
Troubleshooting
Seizure, circling, or sharp weight loss follow neurologic endpoints—do not delay for one more MRI.
| Sign | Cause | Action |
|---|---|---|
| Immediate death / lung bleed | RV or air | Stop; review JoVE; no blind sticks |
| MRI-negative at week 3 | Missed inject, dead cells, wrong subline | Audit flashback; do not jump to the bone-page dose |
| Face/ear mass (carotid) | ECA distribution | Off-model for brain[3] |
| One large intracranial mass (IC route) | Local implant, not metastasis | Do not call it a BCBM cascade |
Monitoring log fields
Neurologic signs outrank the image. No caliper volumes on this page.
Literature case comparison
LV numbers are 231-BR; carotid/intracranial are PDX. Open the PMID for JoVE.
| Paper | Host | Cells | Method | Readout |
|---|---|---|---|---|
| Yoneda 2001[1] | ♀ nude 4 wk | BR 1×10⁵ / 0.1 mL PBS; 28G | LV after bright-red return | BR almost brain-only; BO bone-only; parental bone ± viscera/brain |
| Zhou JoVE 2014[2] | ♀ nude 6–8 wk | 231-BR-GFP 1.75×10⁵ / 100 μL serum-free | US-guided LV | Multifocal MRI ~wk 3; manual success ~50% |
| Liu 2019[3] | ♀ SCID ~8 wk | PDX 1×10⁵ | ICA 50 μL or IC 1–2 μL | BBB transit vs local growth; not an MDA-231 default dose |
| Zhou PLOS 2013[4] | ♀ nude 6–8 wk | 231-BR 2×10⁵ / 100 μL serum-free | US-guided LV | T2 by wk 3; subset enhance by wk 5. Anesthetic doses from that paper are not copied here |
- 01[Ethics] IACUC; seizure/circling/weight-loss endpoints. LV/microsurgical training. RUO. Watch Zhou JoVE[2]. No anesthetic doses here (do not copy the inhalant concentration from the PLOS paper onto this page).
Default is 231-BR LV—not BO from the bone page, not a fat pad. No Matrigel. Anesthetic doses come only from the IACUC SOP.[1][2]
Item Per mouse / study Notes MDA-231BR (or parental control) MC-h133 Start from 1 vial STR/mycoplasma-qualified; log BR vs parental vs BO separately[1] FBS MC100 10% complete medium Regular grade Serum-free DMEM or ice-cold PBS 100 μL/mouse (LV) Zhou PLOS: 100 μL serum-free medium[4]; Yoneda: 0.1 mL PBS[1] 28G insulin syringe (LV) 1 per mouse Yoneda 28G; US-guided method on JoVE[1][2] 33G bent needle (carotid only) Per surgical n Liu PDX: CCA inject, 50 μL; not the MDA-231 default dose[3] Microsyringe (intracranial only) 1–2 μL Liu: 1×10⁵ in 1–2 μL striatum; local growth, not the cascade[3] MRI / optional BLI Shared Multifocal T2 lesions ~week 3; contrast per imaging SOP[2][4] - 02
- 03
- 04[Carotid] Only if BBB transit is required and the surgeon is trained: Liu 1×10⁵ / 50 μL PBS, 33G, ~2 min; those numbers are PDX—pilot first[3].
Dose and host are PDX/SCID—not the MDA-231BR default. Caudal CCA + ECA ligatures send cells via ICA. Watch Figure 1; not a substitute for microsurgical training. No anesthetic doses here.[3]
Check Pass If fail Host Liu: female SCID ~8 wk[3] Do not default to 4-wk nude LV hosts Volume 1×10⁵ / 50 μL PBS; 33G over ~2 min[3] Do not inject the 100 μL LV volume Direction Block caudal backflow; ECA ligature cuts face/ear seeding[3] Face/ear tumor = ECA distribution Readout meaning Hematogenous colonization across the BBB Still not the orthotopic intravasation cascade - 05[Intracranial] 1×10⁵ in 1–2 μL striatum scores local growth—do not call it a metastatic cascade[3].
Liu: 1×10⁵ in 1–2 μL, right striatum. Reliable take—do not extrapolate homing or BBB transit.[3]
Check Pass If fail Volume 1–2 μL, slow 10 μL injures/spills Coordinates 2 mm right of bregma, 2.5 mm deep[3] Wrong coordinates = ventricle or extra-axial What it models Growth in brain parenchyma Use spontaneous orthotopic for cascade; LV/carotid for hematogenous spread - 06[Monitor] Weight and neuro signs; MRI ~week 3. Zhou still sees extracranial lung/bone—log them[2].
Neurologic signs outrank the image. No caliper volumes on this page.
Seizure, circling, or sharp weight loss follow neurologic endpoints—do not delay for one more MRI.
Sign Cause Action Immediate death / lung bleed RV or air Stop; review JoVE; no blind sticks MRI-negative at week 3 Missed inject, dead cells, wrong subline Audit flashback; do not jump to the bone-page dose Face/ear mass (carotid) ECA distribution Off-model for brain[3] One large intracranial mass (IC route) Local implant, not metastasis Do not call it a BCBM cascade - 07[Endpoint] Serial whole-brain sections; screen extracranial sites unless using BR.
- 08[QC] Fix subline source/passage; independent repeats. Do not put BR on the bone page.
Reagents / materials
| Item | Role | Conc. / dose |
|---|---|---|
| MDA-231BR (brain-seeking subline) | LV brain-mets inoculum | 1–2×10⁵ / 100 μL 无血清培养液或 PBS[[1]][[2]][[4]] |
| MRI contrast (optional) | Later BTB permeability | 按影像 SOP；早期灶可增强阴性[[4]] |
| 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
Brain lesion count/volume, MRI/BLI, neurologic signs, whole-brain pathology, optional extracranial mets.
References (PubMed)
- [1]PMID 11499871 — A bone-seeking clone exhibits different biological properties from the MDA-MB-231 parental human breast cancer cells and a brain-seeking clone in vivo and in vitro. J Bone Miner Res (2001)
- [2]PMID 24637963 — Ultrasound imaging-guided intracardiac injection to develop a mouse model of breast cancer brain metastases followed by longitudinal MRI. J Vis Exp (2014)
- [3]PMID 30679540 — Improving orthotopic mouse models of patient-derived breast cancer brain metastases by a modified intracarotid injection method. Sci Rep (2019)
- [4]PMID 23638013 — Longitudinal MRI evaluation of intracranial development and vascular characteristics of breast cancer brain metastases in a mouse model. PLoS One (2013)
Disclaimer: RUO; IACUC required. Optimize by strain and pilot. Red tags mark weak or non-metastatic parental endpoints.