NCI-H69 Cell Line

Brief Introduction
Product List

General Information
Organism	Homo sapiens, human
Cell Line Description	NCI-H69 is a human small cell lung cancer cell line established from metastatic pleural effusion collected from a 55-year-old male. The cells grow as rounded suspension cells that form multicellular aggregates and retain a classic neuroendocrine small cell lung cancer phenotype. NCI-H69 is ASCL1-positive and carries inactivating TP53 and RB1 variants together with a PIK3CA deletion. These features support research on neuroendocrine differentiation, tumor-cell aggregation, growth and survival signaling, apoptosis, chemotherapy response, acquired resistance, radiobiology, three-dimensional culture, and xenograft behavior. The parental line must be distinguished from H69AR, H69V, H69/LX, and other adapted or drug-selected derivatives because these sublines differ in morphology, lineage state, and treatment response.
Cell Type	Small cell lung carcinoma cell
Tissue of Origin	Lung cancer cells recovered from pleural effusion
Disease	Small cell lung carcinoma
Gender	Male
Age	55 years
Population / Ancestry	Caucasian
Morphology	Rounded, lymphoblast-like cells that commonly form floating aggregates
Growth Mode	Suspension with multicellular clustering
Applications	1. Small cell lung cancer biology 2. ASCL1-driven neuroendocrine differentiation 3. TP53- and RB1-deficient tumor research 4. PI3K pathway and PIK3CA-variant studies 5. Cell aggregation and spheroid-like growth 6. Apoptosis and BCL-2 pathway analysis 7. Cytotoxic and targeted-drug screening 8. Chemotherapy response and resistance studies 9. Comparison with variant or drug-resistant H69 derivatives 10. Radiation-response research 11. Invasion, tumorigenicity, and xenograft studies 12. Transcriptomic, proteomic, and genomic profiling
Characteristics
ASCL1 Status	ASCL1-positive. This lineage state distinguishes parental NCI-H69 from the ASCL1-negative variant derivative H69AR.
Neuroendocrine Markers	Classic small cell lung cancer cultures express elevated L-dopa decarboxylase, bombesin-like immunoreactivity, neuron-specific enolase, and the brain isozyme of creatine kinase.
TP53 Variant	TP53 c.511G>T, producing p.Glu171Ter
RB1 Variant	RB1 c.2242G>T, producing p.Glu748Ter
PIK3CA Variant	PIK3CA c.317_325delGGCAACCGT, producing p.Gly106_Arg108del
Microsatellite Status	Microsatellite stable
Karyotype	This is an aneuploid human male cell line. In this hypotetraploid cell line, various normal chromosomes are observed in monosomic or disomic states; however, translocations and deletions involving numerous chromosomes are also observed, and these chromosomal rearrangements appear to be stable and typically occur in paired configurations.
Tumorigenic	Yes, in nude mice
Mycoplasma Test	Negative
Culture Conditions and Handling
Culture Conditions	37°C in a humidified atmosphere containing 5% CO₂ in air
Culture Surface	Standard tissue-culture flasks; surface coating is not required for routine suspension propagation
Recommended Cell-Density Range	Maintain approximately 6 × 10⁴ to 5 × 10⁵ viable cells/mL
Recommended Split Ratio	1:2 to 1:4
Medium Renewal	Add fresh complete medium or perform a complete medium exchange every 2–3 days according to viable-cell density and culture condition
Routine Maintenance	Maintain the suspension by adding fresh complete medium as cell density increases. For complete medium replacement, collect the cells by gentle centrifugation, remove spent medium, and resuspend at the validated starting density.
Subculturing	1. Mix the flask gently to distribute floating aggregates without vigorous shear. 2. Examine aggregate size, debris, and medium color microscopically. 3. Determine viable-cell density using a reproducible sampling and dissociation method. 4. Dilute directly with fresh complete medium or collect the cells for complete medium replacement. 5. If centrifuging, use a validated low-speed setting and remove the supernatant without disturbing the pellet. 6. Resuspend gently and seed new vessels at a 1:2 to 1:4 ratio, targeting approximately 6 × 10⁴ to 5 × 10⁵ viable cells/mL. 7. Return cultures to 37°C and 5% CO₂ and reassess density and aggregation within 2–3 days.
Thawing	Thaw rapidly with gentle agitation in a 37°C water bath for approximately 2 minutes. Disinfect the vial exterior and transfer the contents aseptically into pre-warmed complete medium.
Cryopreservation Medium	Complete growth medium supplemented with 5% DMSO
Long-Term Storage	Below −130°C, preferably in the vapor phase of liquid nitrogen; −70°C storage is not suitable for continued preservation
Post-Thaw Monitoring	Track viable-cell density, aggregate formation, debris, medium condition, and recovery trend. Delay experiments until growth is stable and the culture meets predefined identity and quality criteria.
Biosafety	Handle as human-derived research material under Biosafety Level 1 or the locally assigned equivalent. Apply site-specific risk assessment, aseptic technique, appropriate personal protective equipment, decontamination, and waste procedures.
Handling Notes	Record passage number, viable-cell density, aggregate appearance, split ratio, medium and serum lot, feeding schedule, and time after seeding. Label parental NCI-H69 separately from every resistant, variant, reporter-labeled, gene-edited, or otherwise adapted derivative.

Cat.No.	Product Name	Price
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CSC-RO01528	OVAL Stable Cell Line - NCI-H69	Inquiry
CSC-RO01781	GFP/OVAL Stable Cell Line - NCI-H69	Inquiry
CSC-RO02034	Luc/OVAL Stable Cell Line - NCI-H69	Inquiry
CSC-RO02287	GFP/Luc/OVAL Stable Cell Line - NCI-H69	Inquiry
CSC-RR01263	Luciferase Reporter Cell Line - NCI-H69	Inquiry

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

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