HEp-2 Cells
Introduction to Hep-2 cells
Description | The HEp-2 cell line, originally believed to be derived from laryngeal cancer cells, was later identified through DNA fingerprinting and the presence of HeLa marker chromosomes as being contaminated with HeLa cells, a cell line that was derived from cervical cancer. Despite this, the HEp-2 cell line remains extensively utilized in indirect immunofluorescence to detect antinuclear antibodies (ANAs), which are key in diagnosing conditions like systemic lupus erythematosus and systemic sclerosis. The indirect immunofluorescence assay (IIFA) using HEp-2 cells, which provides clear positive or negative results, is the standard method for testing antinuclear antibodies. This straightforward approach is crucial for diagnosing and classifying different systemic autoimmune diseases. The patterns of autoantibodies observed in indirect immunofluorescence on HEp-2 cells, especially in the context of rheumatology, provide invaluable insights into various rheumatic diseases. Furthermore, the comprehensive review of antigens expressed by HEp-2 human cells under different culture conditions enables the identification of specific ANAs linked to diseases like lupus. In conclusion, while the contamination of cell lines like HEp-2 with HeLa cells has prompted concerns in cancer research about the accuracy and reliability of results and their clinical relevance, the utility of Hep-2 in the detection of antinuclear antibodies and its application across various research disciplines underscore its continued importance. The HEp-2 cell line serves as an essential tool in diagnosing and classifying autoimmune diseases, among other applications. |
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Organism | Human |
Tissue | Larynx |
Disease | Adenocarcinoma |
Applications | In rheumatology, indirect immunofluorescence using HEp-2 cells plays a crucial role in diagnosing autoimmune diseases, including systemic lupus erythematosus and systemic sclerosis |
Synonyms | Hep-2, HEP-2, HEp-2/HeLa, Hep 2, Hep2, HEp2, HEP2, H.Ep.-2, H.Ep. #2, H.Ep. No. 2, Hep II, Human Epidermoid carcinoma #2, Human Epithelioma-2 |
Details of the HEp-2 cell line
Age | 30 years |
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Gender | Female |
Ethnicity | African American |
Morphology | Epithelial-like |
Growth properties | Monolayer, adherent |
Documentation
Citation | HEp-2 (Cytion catalog number 300397) |
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Biosafety level | 1 |
Genetic profile
Isoenzymes | G6PD, A |
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Reverse transcriptase | Negative |
Products | Keratin |
Handling of Hep2 cells
Culture Medium | EMEM, w: 2 mM L-Glutamine, w: 1.5 g/L NaHCO3, w: EBSS, w: 1 mM Sodium pyruvate, w: NEAA (Cytion article number 820100c) |
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Medium supplements | Supplement the medium with 10% FBS |
Passaging solution | Accutase |
Subculturing | Remove the old medium from the adherent cells and wash them with PBS that lacks calcium and magnesium. For T25 flasks, use 3-5 ml of PBS, and for T75 flasks, use 5-10 ml. Then, cover the cells completely with Accutase, using 1-2 ml for T25 flasks and 2.5 ml for T75 flasks. Let the cells incubate at room temperature for 8-10 minutes to detach them. After incubation, gently mix the cells with 10 ml of medium to resuspend them, then centrifuge at 300xg for 3 minutes. Discard the supernatant, resuspend the cells in fresh medium, and transfer them into new flasks that already contain fresh medium. |
Split ratio | A ratio of 1:4 to 1:10 is recommended |
Seeding density | 1 x 10^4 cells/cm^2 |
Fluid renewal | 2 to 3 times per week |
Freezing recovery | After thawing, plate the cells at 5 x 10^4 cells/cm^2 and allow the cells to recover from the freezing process and to adhere for at least 24 hours. |
Freeze medium | CM-1 (Cytion catalog number 800100) or CM-ACF (Cytion catalog number 806100) |
Handling of cryopreserved cultures |
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Quality control
Sterility | Mycoplasma contamination is excluded using both PCR-based assays and luminescence-based mycoplasma detection methods. To ensure there is no bacterial, fungal, or yeast contamination, cell cultures are subjected to daily visual inspections. |
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STR profile |
Amelogenin: x,x
CSF1PO: 9,1
D13S317: 12,13.3
D16S539: 9,1
D5S818: 11,12
D7S820: 8,12
TH01: 7
TPOX: 8,12
vWA: 16,18
D3S1358: 15,18
D21S11: 27,28
D18S51: 16
Penta E: 7,17
Penta D: 8,15
D8S1179: 12,13
FGA: 18,21
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Required products
Key features of Freeze Medium CM-1 include:
Broad Compatibility: Effective for a wide range of cell types, including primary cells, stem cells, and established cell lines.
High Viability: Optimized to maximize post-thaw cell recovery and viability, ensuring reliable experimental outcomes.
Ready-to-Use: Conveniently prepared and sterilized for immediate application, reducing preparation time and risk of contamination.
Enhanced Stability: Maintains consistent performance under standard cryopreservation conditions, ensuring reproducible results.
Long Shelf Life: CM-1 is a serum-containing, ready-to-use cryopreservation medium that can be stored in the refrigerator for up to one year.
Using CM-1 for Freezing Cells
To use CM-1 for freezing both adherent and suspension cells, follow these steps:
For adherent cells, wash and dissociate them from the culture substrate. For suspension cells, proceed directly to the next step.
Count the cells to ensure they are at the proper concentration.
Centrifuge the cells to pellet them, then resuspend in CM-1 freeze medium.
Transfer the resuspended cells into cryovials.
Use a slow-freezing method before transferring the cells to long-term storage.
🥶 Method
🔍 Description
💡 Steps
❄️
Manual Freezing
A step-by-step method involving gradual temperature reduction to ensure cell viability.
1️⃣ Place cells in freeze medium in a 4°C freezer for 40 minutes.
2️⃣ Transfer to a -80°C freezer for 24 hours.
3️⃣ Store cells in liquid nitrogen for long-term preservation.
🧊
Using Mr. Frosty
A convenient device that allows for controlled freezing rates without electrical power.
1️⃣ Prepare cells in cryovials with freeze medium.
2️⃣ Place cryovials in Mr. Frosty container.
3️⃣ Store at -80°C for 24 hours before transferring to liquid nitrogen.
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Controlled-Rate Freezer
A high-precision freezer by Thermo Fisher or other manufacturers designed for controlled temperature reduction.
1️⃣ Program the device to gradually decrease the temperature.
2️⃣ Place prepared cells in the freezer.
3️⃣ After the freezing cycle, transfer cells to liquid nitrogen.
Store the cryovials at temperatures below -130°C or in liquid nitrogen for long-term preservation.
Ingredients
Contains FBS, DMSO, Glucose, Salts
Buffering capacity: pH = 7.2 to 7.6
Cytion’s Freeze Medium CM-1 offers a reliable solution for cryopreservation, ensuring high cell viability and functionality post-thaw for a wide range of research applications.
Phosphate-buffered saline (PBS) is a versatile buffer solution used in many biological and chemical applications, as well as tissue processing. Our PBS solution is formulated with high-quality ingredients to ensure a constant pH during experiments. The osmolarity and ion concentrations of our PBS solution are matched to those of the human body, making it isotonic and non-toxic to most cells.
Composition of our PBS Solution
Our PBS solution is a pH-adjusted blend of ultrapure-grade phosphate buffers and saline solutions. At a 1X working concentration, it contains 137 mM NaCl, 2.7 mM KCl, 8 mM Na2HPO4, and 2 mM KH2PO4. We have chosen this composition based on CSHL protocols and Molecular cloning by Sambrook, which are well-established standards in the research community.
Applications of our PBS Solution
Our PBS solution is ideal for a wide range of applications in biological research. Its isotonic and non-toxic properties make it perfect for substance dilution and cell container rinsing. Our PBS solution with EDTA can also be used to disengage attached and clumped cells. However, it is important to note that divalent metals such as zinc cannot be added to PBS as this may result in precipitation. In such cases, Good's buffers are recommended. Moreover, our PBS solution has been shown to be an acceptable alternative to viral transport medium for the transport and storage of RNA viruses, such as SARS-CoV-2.
Storage of our PBS Solution
Our PBS solution can be stored at room temperature, making it easy to use and access.
To sum up
In summary, our PBS solution is an essential component in many biological and chemical experiments. Its isotonic and non-toxic properties make it suitable for numerous applications, from cell culture to viral transport medium. By choosing our high-quality PBS solution, researchers can optimize their experiments and ensure accurate and reliable results.
Composition
Components
mg/L
Inorganic Salts
Potassium chloride
200,00
Potassium dihydrogen phosphate
200,00
Sodium chloride
8,000.00
di-Sodium hydrogen phosphate anhydrous
1,150.00