Cell Culture / Organoids / Endometrial cancer PDO

Endometrial cancer / disease PDO banking protocol

Patient-derived endometrial epithelial organoids: embed in an endometrial niche (EGF / Noggin / R-spondin / FGF10, A83-01, nicotinamide…; ±E2 for hormone arms), covering cancer and some benign disease, then QC with PAX8 / CK / ERα and run drugs.[1][2]

Use cases

Endometrial PDO / disease organoids—not Ishikawa spheroids and not a full decidualization course.

UseFitReadoutNotes
Endometrial cancer / hyperplasia / endometriosis biobank + drugsRecommendedSubtype fidelity and patient-specific drugs[1]Boretto platform covers cancer and multiple endometrial diseases and can recapitulate lesions in vivo.
Normal endometrium hormone-response contrast (Turco-style)Contrast±E2 morphology / markersSeparate from tumor dishes; lock E2 / serum lots for hormone claims.
Ishikawa spheroids as PDODo not mixSpheroid diameterCell-line spheroids are not PDOs.

In one line

Endometrial clusters → dome → EGF/Rspo/FGF10 niche → brief Y-27632 → mechanical split → PAX8/ER → drugs.

Protocol overview

  1. Ethics + cold chain; log histology, grade, and hormone-receptor status
  2. Trim necrosis; collagenase to clusters; brief RBC lysis (MC427) if bloody
  3. Matrix domes; endometrial niche (EGF, Noggin, R-spondin, FGF10, A83-01, nicotinamide…)[[1]][[2]]
  4. Brief Y-27632; score cystic budding
  5. Feed every 2–3 days; mechanical split; bank cancer vs benign / endometriosis separately
  6. PAX8, CK8/18, EpCAM, ERα by study; mycoplasma
  7. Hormone arms: serum-free or CSS±E2±progesterone; separate from default expansion dishes
  8. Drugs with vehicle; in vivo uses transplant SOPs—not this page

Step 1 · Specimen processing

  1. Log stage / molecular class if known and hormone receptors.
  2. Collagenase to clusters; brief RBC lysis (MC427) if bloody.
  3. Separate instruments and dishes for cancer, hyperplasia, and endometriosis.

Steps 2–3 · Niche and hormones

  1. Start with a full endometrial recipe[1][2].
  2. Hormone manipulation (±E2 or CSS±E2) on separate dishes—do not mix with default expansion lots.
  3. If stroma overtakes, raise A83-01.

Steps 4–5 · QC and drugs

  1. PAX8, CK8/18, EpCAM, ERα; mycoplasma.
  2. Include vehicle; interpret vs clinical regimens[1].
  3. In vivo confirmation uses transplant models—not this page.

Niche reagents (titrate)

Literature starting points—not on-site prescribed doses.

ComponentRoleOn-site
EGF + Noggin + R-spondin + FGF10Endometrial core[1][2]Open →
A83-01 / 烟酰胺Human epithelial stabilityOpen →
雌二醇 E2（激素臂）Optional in expansion; hormone arms on separate dishesOpen →
孕酮（激素臂）Often with E2; titrate on separate dishesOpen →
Y-27632 短期Seeding windowOpen →
红细胞裂解液Brief RBC lysis if bloodyOpen →
胶原酶 II 型Digest tissue/tumor to epithelial clustersOpen →
Dispase IIGentle dissociation with collagenaseOpen →
类器官冻存液Banking cryopreservationOpen →

Small molecules for this SOP

Deep-links to catalog SKUs; all prices on request. Growth factors (EGF/Noggin/R-spondin, etc.) are proteins—see that category.

CompoundSKURolePrice
Y-27632 2HClMCK1042ROCK block; brief survival boost after seed/splitOn request
SB 202190MCK1279p38 block; human epithelial organoid stabilityOn request
A83-01MCK1825ALK4/5/7; limit stromal overgrowthOn request
17β-EstradiolMCK1835Hormone arm for ER+ breast organoidsOn request
ProgesteroneMCK1838Breast / endometrial hormone arms; with E2 on separate dishesOn request
NicotinamideMCK1836Common in human epithelial organoid mediaOn request
Inquire for this listAll small molecules

Proteins / peptides for this SOP

Growth factors and gastrin deep-linked by SKU; all prices on request. Activity/reconstitution per lot COA.

ProductSKURoleTypePrice
Recombinant Human EGFMGF30001Core epithelial growth factorProteinOn request
Recombinant Human NogginMGF30002BMP antagonist; ENR / PDO niche; dual-SMAD neural inductionProteinOn request
Recombinant Human R-spondin-1MGF30003Wnt amplifier; long-term expansionProteinOn request
Recombinant Human FGF10MGF30007Prostate / pancreas / breast / lung / gastric / ovarian / liver / endometrial / kidney nicheProteinOn request
Inquire for this listProteins / enzymes / peptides

QC antibodies for this SOP

Common IHC/IF markers before banking or drugs; deep-linked catalog antibodies. Dilution/species per datasheet.

MarkerProductSKURole
CK8Cytokeratin 8 AntibodyMCAB5383Luminal / epithelial lineage
CK18Cytokeratin 18 AntibodyMCAB5028Luminal epithelium; breast / prostate / ovarian / endometrial
ERαER alpha AntibodyMCAB5041Breast / endometrial subtype vs clinical pathology
PRProgesterone Receptor AntibodyMCAB5590Breast / endometrial hormone-receptor QC
EpCAMEPCAM AntibodyMCAB5377Pan-epithelial; multi-tissue PDO
E-CadherinE Cadherin AntibodyMCAB1155Epithelial integrity / EMT contrast
PAX8PAX8 AntibodyMCAB5911Ovarian / Müllerian / endometrial / renal epithelial QC
Inquire for this listAntibodies

Culture supplements for this SOP

Existing cell-culture SKUs (B-27 / N-2 / glutamine / antibiotics); titrate per insert and literature.

ProductSKURole
Advanced DMEM/F12 MediumMC410Serum-free organoid basal medium
B-27 Supplement (50×)MC317Serum-free basal supplement for epithelial / neural organoids
Stable glutamine (Ala-Gln, 200 mM)MC301Glutamine source for Advanced DMEM/F12 systems
Penicillin-Streptomycin (100×)MC306Early banking sterility; consider withdrawal after lines stabilize
Organoid Basement Membrane Extract (BME)MC4113D dome / sandwich embedding
Collagenase Type IIMC412Digest tissue/tumor to epithelial clusters
Dispase IIMC413Gentle dissociation with collagenase
Accutase Cell Detachment SolutionMC414Optional gentle enzymatic passaging
Organoid Cryopreservation MediumMC415Banking cryopreservation
Mycoplasma Detection KitMC416Pre-banking sterility QC
Charcoal-Stripped FBS (CSS)MC419Serum + hormone manipulation only; not for default serum-free banking
HEPES Buffer (1 M)MC305Organoid medium buffering; add to final concentration per recipe
Collagenase Type IVMC421Dissociation alternative; contrast with Type II
Organoid Primary Antimicrobial (Primocin-class)MC423Early primary / PDO / PSC banking contamination control
Ultra-Low Attachment 96-Well PlateMC425EB formation / spheroids / drug plates
U-Bottom 96-Well PlateMC430Spheroid settling / co-culture plates; complements ULA
RBC Lysis BufferMC427RBC removal after bloody digests
Inquire for this listCulture supplements

References

  1. [1] Patient-derived organoids from endometrial disease capture clinical heterogeneity and are amenable to drug screening. Nat Cell Biol. 2019. PMID 31371824
  2. [2] Long-term, hormone-responsive organoid cultures of human endometrium in a chemically defined medium. Nat Cell Biol. 2017. PMID 28319085

Related

Other SOPs

Disclaimer: Research use only (RUO). Not clinical guidance or an institutional SOP. Human tissue requires ethics approval. Inline [n] maps to the reference list. Titrate concentrations and digest times locally.