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COVID-19 Spike (S) Protein / HeLa Cell Line

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Stable HeLa cell line constitutively expressing full-length SARS-CoV-2 Spike (S) glycoprotein with a C-terminal eGFP tag under CMV promoter. Greater than 85% Spike/eGFP co-expression confirmed by flow cytometry. ACE2 binding validated by recombinant ACE2-Fc functional assay. Suitable for viral entry assays, SARS-CoV-2 neutralization studies, anti-Spike antibody screening, and ACE2/Spike interaction research. Does not produce infectious virus. BSL-2.s.

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
Return & Refund Policy

Internal Reference: KCL-ENG-202

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

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

KCL-ENG-202 is a stable, adherent HeLa cell line engineered to constitutively express the full-length SARS-CoV-2 Spike (S) glycoprotein under a CMV promoter. The Spike protein carries a C-terminal enhanced GFP (eGFP) tag for direct fluorescence visualization and flow cytometry detection without requiring anti-Spike antibody staining in every assay. The line is maintained under Puromycin selection to ensure stable transgene retention.

Greater than 85% of cells co-express Spike and eGFP as confirmed by flow cytometry. Surface Spike expression is validated by anti-Spike antibody staining (92% positive; MFI greater than 10,000), and functional receptor interaction is confirmed by dose-dependent binding of recombinant human ACE2-Fc. Western blot shows the expected full-length band at approximately 180 kDa. Use within 10 passages post-thaw to ensure consistent stable expression.

This cell line does not produce infectious SARS-CoV-2 virus. It expresses Spike protein only and is suitable for BSL-2 work. All lots are quality tested prior to release for Spike/GFP co-expression, viability, mycoplasma, and sterility.

Recommended Medium

DMEM supplemented with 10% heat-inactivated FBS and 1% Pen/Strep. Maintain under Puromycin selection at 1 μg/mL for routine culture. Omit Puromycin during experimental assays as needed. Subculture at 80 to 90% confluence using 0.25% Trypsin-EDTA; split 1:4 to 1:6 every 2 to 3 days.

Applications
SARS-CoV-2 Viral Entry Assays Neutralization Studies Anti-Spike Antibody Screening ACE2 / Spike Interaction Studies Spike Protein Immunology Research Flow Cytometry Spike Positive Control Fluorescence-Based Co-culture Assays Antibody Internalization Studies
Characterization

All lots are quality tested prior to release. Expression validation is performed at passage 4. The following data is representative of lot release testing.

Expression & Identity
Parameter Result Specification
SARS-CoV-2 Spike surface expression ✓ Positive 92% by flow cytometry (anti-Spike clone 1A9); MFI >10,000
eGFP fluorescence ✓ Positive 89% by flow cytometry (GFP channel); C-terminal co-localized with Spike
Spike / eGFP co-expression ✓ Confirmed >85% double-positive of live cells
Spike protein (Western blot) ✓ Confirmed Band at ~180 kDa (full-length); anti-S1/RBD antibody; reducing conditions
ACE2 binding (functional) ✓ Positive Dose-dependent binding of recombinant human ACE2-Fc; ≥1-log shift over parental HeLa
Post-thaw viability ✓ Pass >90% by Trypan Blue exclusion
Passage number at freeze ✓ Confirmed Passage 1
Quality Control
Test Result Method
Mycoplasma ✗ Negative PCR (EP 2.6.7)
Sterility — Bacteria ✗ Negative 14-day culture
Sterility — Fungi & Yeast ✗ Negative 14-day culture

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

Receipt Protocol

KCL-ENG-202 is shipped as a cryopreserved vial on dry ice. These are adherent HeLa cells. Follow the steps below upon receipt. Process within 48 hours of delivery.

❄  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. Remove when a small ice crystal remains.
2
Transfer cells dropwise into 10 mL of pre-warmed DMEM + 10% FBS (no Puromycin). Centrifuge at 200 × g for 5 minutes. Aspirate supernatant to remove DMSO.
3
Resuspend in complete growth medium + 1 μg/mL Puromycin. Seed into a T-25 flask at 37°C, 5% CO₂.
4
After 24 hours, replace medium to remove dead cells and debris. Continue culture under Puromycin selection.
⚠ Do not leave thawed cells without medium — immediate culture is required. Do not store thawed cells at 4°C. Use within 10 passages post-thaw to ensure stable Spike/eGFP expression.
Subculture Guidelines

Subculture at 80 to 90% confluence using 0.25% Trypsin-EDTA. Split 1:4 to 1:6 every 2 to 3 days. Maintain under Puromycin selection at 1 μg/mL. Verify Spike and eGFP co-expression by flow cytometry periodically, particularly before critical experiments. Re-establish from frozen stock after passage 10. Full protocols available on the Kulture Scientific protocol page.

Product Catalog

KCL-ENG-202 is available as a cryopreserved vial containing at least 5×10&sup5; viable cells in cryoprotective medium.

Product Cat. No. Format Cell Number
COVID-19 Spike (S) Protein / HeLa Cell Line KCL-ENG-202 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); cells are stable indefinitely under correct cryogenic conditions
  • 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 Spike/eGFP co-expression stability; thaw only the quantity needed for immediate use
  • Shipping condition — cryopreserved vials are shipped on dry ice; process within 48 hours of delivery and transfer to liquid nitrogen storage immediately upon receipt
  • Post-thaw passage limit — use within 10 passages post-thaw; re-establish from frozen stock after passage 10 to maintain stable expression
  • DMSO content — cryoprotective medium contains DMSO; wear nitrile gloves during thaw procedures and avoid skin contact
  • Certificate of Analysis — lot-specific QC documentation including Spike and GFP flow cytometry data and viability results available upon request
Health & Safety Information
⚠ WARNING! This product is of human origin. Handle all human-derived biological materials using Universal Precautions in accordance with local BSL-2 regulations. Appropriate PPE must be worn at all times, including gloves, lab coat, and eye protection. All waste must be autoclaved or treated with 10% bleach prior to disposal. Safety Data Sheet available at kulturesci.com.
Regulatory Status

For research purposes only. Not for use in diagnostic or therapeutic procedures. Not for in vivo use without further validation. This product has not been tested or validated for clinical use. Not for direct administration into humans or animals.

Biosafety Level

This cell line should be handled under Biosafety Level 2 (BSL-2) conditions. This product does not contain or produce infectious SARS-CoV-2 virus. BSL-2 is required based on the following:

  • Human-derived origin — HeLa cells are derived from human cervical adenocarcinoma tissue; all human-derived cell lines require BSL-2 containment under Universal Precautions
  • SARS-CoV-2 Spike transgene — expresses the viral Spike glycoprotein; does not contain the full SARS-CoV-2 genome and cannot produce infectious virus; not a select agent
  • Recombinant cell line — stable lentiviral or plasmid integration; standard BSL-2 containment for recombinant human cell lines applies

Users must consult their institutional biosafety committee (IBC) prior to use. Individual institutions may impose additional requirements based on local policy regarding SARS-CoV-2-related research materials.

Biosafety Considerations
  • Human-derived material — treat as potentially infectious; work in a certified biosafety cabinet; follow BSL-2 guidelines at all times
  • Spike protein expression — this cell line expresses SARS-CoV-2 Spike glycoprotein on the cell surface; it cannot produce infectious virus but should be treated with appropriate containment
  • Spill response — cover with 10% bleach, allow 20 minutes contact time, absorb, and decontaminate the surface; document all spill incidents per institutional protocol
  • DMSO (cryoprotectant) — skin-penetrating solvent; wear nitrile gloves at all times during thawing
  • Waste disposal — autoclave all cell culture waste or treat with 10% bleach prior to disposal in accordance with local biohazardous waste regulations
Frequently Asked Questions
Does this cell line produce infectious SARS-CoV-2?
No. KCL-ENG-202 expresses only the Spike (S) glycoprotein and does not contain the full SARS-CoV-2 genome. It cannot produce infectious virus. This product is not a select agent. However, BSL-2 practices are still required because the cells are of human origin and express a viral surface glycoprotein. Consult your institutional biosafety committee prior to use.
Why does the Spike protein have a C-terminal eGFP tag?
The eGFP tag serves as a built-in reporter for direct visualization of Spike-expressing cells by fluorescence microscopy or flow cytometry without requiring anti-Spike antibody staining in every experiment. It also enables real-time monitoring of expression levels during culture and provides a convenient readout for co-culture and cell tracking assays. The tag is at the C-terminus of the Spike protein, which preserves the native N-terminal signal peptide and receptor-binding domain (RBD) orientation required for ACE2 interaction.
Has functional ACE2 binding been confirmed?
Yes. Functional receptor interaction has been confirmed by incubation with recombinant human ACE2-Fc followed by anti-Fc-PE detection, showing a shift of at least 1 log over parental HeLa cells. This confirms that the expressed Spike protein maintains a conformation capable of engaging its primary receptor, making this line suitable for receptor binding and entry inhibition assays.
How many passages can I use this line for?
Use within 10 passages post-thaw. Beyond this point, Spike and eGFP co-expression may decline without consistent selection pressure. Always maintain under Puromycin selection at 1 μg/mL and verify expression by flow cytometry periodically. Re-establish culture from a new frozen vial after passage 10 to ensure assay consistency.
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.
Where can I find the Certificate of Analysis for my lot?
Certificates of Analysis are available upon request. Contact Kulture Scientific at info@kulturesci.com with the lot number printed on your vial label to receive the corresponding QC documentation including flow cytometry data for Spike and GFP co-expression, viability, and sterility results.