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CHO-K1 / CD19 & GFP Expression Stable Cell Line

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Genetically engineered CHO-K1 stable cell line constitutively co-expressing full-length human CD19 and Green Fluorescent Protein (GFP). Greater than 90% CD19 and GFP co-expression confirmed by flow cytometry through passage 15. Non-human CHO-K1 background eliminates endogenous human surface marker interference — ideal for anti-CD19 antibody screening, CAR-T cell cytotoxicity assays, fluorescence-based killing assays, and live-cell co-culture imaging. BSL-1 classification simplifies handling compared to human-derived CD19 cell lines.

Storage and Shipping: Cells are shipped as a frozen vial on dry ice. 

Upon Receipt: Frozen Vial: Transfer immediately to liquid nitrogen for long-term storage. 

⚠️ FOR RESEARCH USE ONLY This product is intended for laboratory research purposes only and is not suitable for human or veterinary diagnostic or therapeutic use.
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For Research Use Only
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Internal Reference: KCL-ENG-201

Description
Characterization
Receipt Protocol
Catalog
Storage & Handling
Safety
FAQs
Product Description

⚠ FOR RESEARCH USE ONLY — Not for diagnostic or therapeutic procedures

KCL-ENG-201 is a genetically engineered CHO-K1 (Chinese Hamster Ovary) cell line that constitutively and stably co-expresses full-length human CD19 and Green Fluorescent Protein (GFP). The dual-expression system uses a bicistronic or separate promoter design to ensure high, coordinated expression of both transgenes from a single stably integrated construct.

Greater than 90% of cells co-express CD19 and GFP through passage 15 as verified by flow cytometry, with CD19 expression confirmed at greater than 95% positive (MFI at least 1.5-fold over isotype control) and GFP fluorescence confirmed at greater than 95% positive (Ex/Em 488/510 nm). The CHO-K1 background is non-human in origin, providing a clean, low-background host for CD19 display without endogenous human surface markers that could confound assay readouts.

This line is particularly well suited to fluorescence-based CAR-T cytotoxicity assays, where GFP enables real-time or endpoint target cell quantification without the need for additional labeling. All lots are quality tested prior to release for co-expression, viability, mycoplasma, and sterility.

Recommended Medium

Ham's F-12K supplemented with 10% heat-inactivated FBS and 1% Pen/Strep. For long-term maintenance of stable expression, selection antibiotic may be applied for the first 2 passages after thaw (Puromycin at 5 μg/mL or Neomycin as appropriate — confirm selection marker with Certificate of Analysis for your lot). Subculture at 80 to 90% confluence using 0.25% Trypsin-EDTA.

Applications
CAR-T Cell Cytotoxicity Assays Fluorescence-Based Killing Assays Anti-CD19 Antibody Screening Antibody Internalization Studies Flow Cytometry CD19 Positive Control Live-Cell Fluorescence Imaging Co-culture Assays Receptor-Ligand Interaction Studies
Characterization

All lots are quality tested prior to release. Co-expression of CD19 and GFP is confirmed by flow cytometry for each production lot.

Expression & Identity
Parameter Result Specification
Human CD19 expression ✓ Positive >95% by flow cytometry; MFI ≥1.5× isotype control
GFP fluorescence ✓ Positive >95% by flow cytometry; Ex/Em 488/510 nm
CD19 / GFP co-expression ✓ Stable >90% co-expression through passage 15
Post-thaw viability ✓ Pass ≥85% by Trypan Blue; 88% average across thaw testing (n=3)
Passage number at freeze ✓ Confirmed P2 to P3
Parental cell line ✓ Confirmed CHO-K1; adherent, epithelial-like; ~22 to 24 hr doubling time
Quality Control
Test Result
Mycoplasma ✗ Negative
Sterility — Bacteria ✗ Negative
Sterility — Fungi ✗ Negative

Certificates of Analysis with lot-specific QC data including flow cytometry histograms for CD19 and GFP co-expression are available upon request. Contact Kulture Scientific with your lot number.

Receipt Protocol

KCL-ENG-201 is shipped as a cryopreserved vial on dry ice. These are adherent cells — follow the steps below upon receipt.

❄  Immediate receipt (same day)
1
Inspect the vial upon receipt. Confirm dry ice is present and the vial cap is intact. Do not allow cells to thaw.
2
Transfer immediately to liquid nitrogen vapor phase (−150°C to −196°C) for long-term storage.
⚠ Do not allow cells to thaw during transit or before intended use. Do not store at −80°C or −20°C.
⚗  Thawing & recovery
1
Thaw vial rapidly in a 37°C water bath with gentle agitation (under 2 minutes). Remove when a small ice crystal remains.
2
Transfer cells dropwise into 9 mL of pre-warmed Ham's F-12K + 10% FBS (no selection antibiotic). Centrifuge at 200 × g for 5 minutes.
3
Aspirate supernatant and resuspend in 10 mL complete growth medium. Transfer to a T-25 flask and incubate at 37°C, 5% CO₂, 95% humidity.
4
After 24 hours, replace medium to remove dead cells and debris. Selection antibiotic (e.g., Puromycin at 5 μg/mL) may be added at this point if desired for the first 2 passages.
⚠ Do not add selection antibiotic immediately at thaw — allow 24 hours recovery first. Confirm the correct selection marker for your lot with the Certificate of Analysis before adding antibiotic.
Subculture Guidelines

Subculture at 80 to 90% confluence using 0.25% Trypsin-EDTA. Split 1:4 to 1:6 every 2 to 3 days. Doubling time is approximately 22 to 24 hours. For long-term culture beyond passage 15, verify co-expression by flow cytometry and consider applying selection antibiotic periodically to maintain stable transgene expression. GFP signal may be affected by fixation protocols — use live-cell imaging or 4% PFA-optimized fixation if downstream immunostaining is required.

Product Catalog

KCL-ENG-201 is available as a cryopreserved vial containing at least 5×10&sup5; viable cells in cryoprotective medium.

Product Cat. No. Format Cell Number
CHO-K1 / CD19 & GFP Expression Stable Cell Line KCL-ENG-201 Cryopreserved Vial ≥5×105 cells/vial
Storage & Handling
❄ Store in Liquid Nitrogen ◾ Ships on Dry Ice ⚠ Do Not Store at −20°C or −80°C
  • Storage — store in liquid nitrogen vapor phase (−150°C to −196°C); this is the only recommended storage condition for cryopreserved cells
  • Do not store at −80°C or −20°C — neither temperature is suitable for long-term cell storage; ice crystal formation at these temperatures will irreversibly damage cell membranes and compromise viability
  • Temperature sensitivity — minimize time outside cryogenic storage; cells are sensitive to temperature fluctuations
  • Freeze-thaw cycles — repeated freeze-thaw cycles compromise viability and co-expression stability; thaw only the quantity needed for immediate use
  • Stability — cells stored in liquid nitrogen are stable indefinitely when maintained under correct cryogenic conditions
  • Shipping condition — cryopreserved vials are shipped on dry ice; inspect packaging upon receipt and transfer to liquid nitrogen storage immediately
  • DMSO content — cryoprotective medium contains DMSO; wear nitrile gloves during thaw procedures and avoid skin contact
  • GFP stability — GFP signal is stable in live cells; verify fluorescence by flow cytometry after thaw. For extended culture beyond passage 15, confirm co-expression periodically
  • Certificate of Analysis — lot-specific QC documentation including CD19 and GFP flow cytometry data and selection marker confirmation available upon request
Health & Safety Information
✓ BSL-1 Product. CHO-K1 cells are of non-human origin and this cell line is classified as Biosafety Level 1. Standard laboratory precautions apply. Wear gloves, lab coat, and safety glasses. Refer to your institutional guidelines for proper handling and disposal of recombinant cell lines. Safety Data Sheet available at kulturesci.com.
Regulatory Status

For research purposes only. Not for use in diagnostic or therapeutic procedures. This product has not been tested or validated for clinical use. Not for direct administration into humans or animals.

Biosafety Level

This cell line is classified as Biosafety Level 1 (BSL-1). CHO-K1 cells are derived from Chinese Hamster Ovary tissue and are not of human origin. The expressed human CD19 transgene is a non-pathogenic surface protein. Standard BSL-1 containment is appropriate for routine culture of this line.

Users should confirm the biosafety classification with their institutional biosafety committee (IBC) prior to use, particularly if the line will be used in co-culture experiments with human-derived immune cells, which may require BSL-2 conditions for the co-culture component.

Biosafety Considerations
  • Recombinant cell line — dispose of all cell culture waste by autoclaving or chemical disinfection (10% bleach) in accordance with institutional guidelines for genetically modified organisms
  • DMSO (cryoprotectant) — skin-penetrating solvent; wear nitrile gloves at all times during thawing
  • GFP — derived from Aequorea victoria; non-toxic, non-pathogenic; no additional containment required beyond standard recombinant cell line handling
  • Co-culture with human immune cells — if this line is used in co-culture with human-derived cells (e.g., CAR-T cells), the co-culture should be handled under BSL-2 conditions appropriate for the human-derived component
Frequently Asked Questions
Why use a CHO-K1 background instead of HEK293 for CD19 expression?
CHO-K1 cells are non-human in origin and do not express human surface markers endogenously, providing a cleaner background for CD19 display assays where human-specific antibodies might cross-react with HEK293 surface proteins. CHO-K1 is also classified as BSL-1, which simplifies handling requirements compared to human-derived lines. The trade-off is that CHO cells lack human post-translational modification machinery, so CD19 glycosylation may differ slightly from native human expression.
Why does this line express GFP alongside CD19?
The GFP co-expression serves as a built-in viability and target cell reporter for fluorescence-based assays. In CAR-T cytotoxicity assays, loss of GFP signal directly quantifies target cell killing without requiring additional labeling steps or fixation. GFP also provides a convenient positive control for flow cytometry panel setup and a tracer for live-cell co-culture imaging.
Does fixation affect the GFP signal?
Yes. Standard aldehyde-based fixatives can quench or alter GFP fluorescence. For best results, use live-cell detection by flow cytometry or fluorescence microscopy. If fixation is required, optimize with 4% paraformaldehyde (PFA) at reduced concentration or shortened exposure time. Avoid methanol-based fixation protocols, which typically abolish GFP signal entirely.
How do I confirm which selection marker my lot uses?
The selection marker (Puromycin or Neomycin/G418) varies by lot. Confirm the correct selection antibiotic and working concentration from the Certificate of Analysis for your specific lot number. Contact Kulture Scientific at info@kulturesci.com with your lot number to request CoA documentation.
Can I store cells at −80°C?
No. Neither −80°C nor −20°C is suitable for long-term cell storage. Ice crystal formation at these temperatures will irreversibly damage cell membranes and compromise viability. Cells must be stored in liquid nitrogen vapor phase (−150°C to −196°C), where they are stable indefinitely.
How long will co-expression remain stable?
Co-expression of CD19 and GFP has been validated as stable through passage 15. Beyond this point, expression may decline without selection pressure. For extended culture, apply selection antibiotic periodically (every 3 to 4 passages) and verify co-expression by flow cytometry before critical experiments.
Is this product suitable for clinical or therapeutic use?
No. KCL-ENG-201 is for research use only and is not validated for diagnostic or therapeutic procedures. Do not administer to humans or animals.