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Primary Bronchial/Tracheal Epithelial Cells; Asthma (ATCC® PCS-300-011™)

Organism: Homo sapiens, human  /  Tissue: respiratory tract  /  Cell Type: epithelial

Permits and Restrictions

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Organism Homo sapiens, human
Tissue respiratory tract
Cell Type epithelial
Morphology epithelial, packed cuboidal morphology
Growth Properties adherent
Biosafety Level 1

[These primary cells are not known to harbor an agent recognized to cause disease in healthy adult humans. Handle as a potentially biohazardous material under at least Biosafety Level 1 containment. Cells derived from primate lymphoid tissue may fall under the regulations of 29 CFR 1910.1030 Bloodborne Pathogens.  

ATCC recommends that appropriate safety procedures be used when handling all primary cells and cell lines, especially those derived from human or other primate material. Detailed discussions of laboratory safety procedures are provided in Laboratory Safety: Principles and Practice, 2nd ed. (ASM Press, Washington, DC) (Fleming et al., 1995) and Caputo, J.L. Biosafety procedures in cell culture. (1988) J. Tissue Culture Methods 11:223.

Appropriate safety procedures should always be used with this material. Laboratory safety is discussed in the following publication: Biosafety in Microbiological and Biomedical Laboratories, 5th ed. HHS Publication No. (CDC) 93-8395. U.S. Department of Health and Human Services, Centers for Disease Control and Prevention. Washington DC: U.S. Government Printing Office; 2007. The entire text is available online at http://www.cdc.gov/biosafety/publications/bmbl5/index.htm.]

Human Material Precaution 

All tissues used for isolation are obtained under informed consent and conform to HIPAA standards to protect the privacy of the donor’s personal health information. It is best to use caution when handling any human cells. We recommend that all human cells be accorded the same level of biosafety consideration as cells known to carry HIV. With infectious virus assays or viral antigen assays, even a negative test result may leave open the possible existence of a latent viral genome.


Biosafety classification is based on U.S. Public Health Service Guidelines, it is the responsibility of the customer to ensure that their facilities comply with biosafety regulations for their own country.
Disease asthma
Gender batch-specific
Ethnicity batch-specific
Applications
Microbial (e.g., viral, bacterial) infection and pathogenesis; airway inflammation and wound healing; asthma; pulmonary fibrosis, chronic obstructive pulmonary disease; chronic bronchitis; toxicology/other testing of pharmaceuticals.
Product Format frozen 1.0 mL
Storage Conditions liquid nitrogen vapor phase
Comments Due to limited availability of appropriate donor material, product inventory may not be available at all times.
Complete Growth Medium
  1. Obtain one Bronchial Epithelial Growth Kit (ATCC® PCS-300-040) from the freezer; make sure that the caps of all components are tight.
  2. Thaw the components of the growth kit just prior to adding them to the basal medium. If the growth kit contains L-glutamine, warm the L-glutamine component in a 37°C water bath and shake to dissolve any precipitates prior to adding to the basal medium.
  3. Obtain one bottle of Airway Cell Basal Medium (485 mL) from cold storage.
  4. Decontaminate the external surfaces of all growth kit component vials and the basal medium bottle by spraying them with 70% ethanol.
  5. Using aseptic technique and working in a laminar flow hood or biosafety cabinet, transfer the indicated volume of each growth kit component, as indicated in Table 1 or 2, to the bottle of basal medium using a separate sterile pipette for each transfer.

Table 1. When using the Bronchial Epithelial Cell Growth Kit (ATCC® PCS-300-040), add the indicated volume for each component: 

Component                     

Volume

Final Concentration

HLL Supplement

1.25 mL

HSA 500 mg/mL

Linoleic Acid 0.6 mM

Lecithin 0.6 mg/mL

L-Glutamine

15 mL

6 mM

Extract P

2.0 mL

0.4%

Airway Epithelial Cell Supplement

5.0 mL

Epinephrine 1.0 mM

Transferrin 5 mg/mL

T3 10 nM

Hydrocortisone 5 mg/mL

rh EGF 5 ng/mL

rh Insulin 5 mg/mL

 

 

Antimicrobials and phenol red are not required for proliferation but may be added if desired. The recommended volume of each optional component to be added to the complete media is summarized in Table 2.

 

Table 2. Addition of Antimicrobials/Antibiotics and Phenol Red (Optional)

Component

Volume

Final Concentration

Gentamicin-Amphotericin B Solution

0.5 mL

Gentamicin: 10 µg/mL

Amphotericin B: 0.25 µg/mL

Penicillin-Streptomycin-Amphotericin B Solution

0.5 mL

Penicillin: 10 Units/mL

Streptomycin: 10 µg/mL

Amphotericin B: 25 ng/mL

Phenol Red

0.5 mL

33 µM
  1. Tightly cap the bottle of complete growth medium and swirl the contents gently to assure a homogeneous solution. Do not shake forcefully to avoid foaming. Label and date the bottle.
  2. Complete growth media should be stored in the dark at 2°C to 8°C (do not freeze). When stored under these conditions, complete media is stable for 30 days. 
Subculturing
1. Passage normal small airway cells when the culture has reached approximately 70% to 80% confluence.
2. Warm both the Trypsin-EDTA for Primary Cells (ATCC PCS-999-003) and the Trypsin Neutralizing Solution (ATCC PCS-999-004) to room temperature prior to dissociation. Warm the complete growth medium to 37°C prior to use with the cells.
3. For each flask, carefully aspirate the spent media without disturbing the monolayer.
4. Rinse the cell layer one time with 3 to 5 mL D-PBS (ATCC 30-2200) to remove residual medium.
5. Add pre-warmed trypsin-EDTA solution (1 to 2 mL for every 25 cm2) to each flask.
6. Gently rock each flask to ensure complete coverage of the trypsin-EDTA solution over the cells, and then aspirate the excess fluid off of the monolayer.
7. Observe the cells under the microscope. When the cells pull away from each other and round up (typically within 1 to 3 minutes), remove the flask from the microscope and gently tap it from several sides to promote detachment of the cells from the flask surface.
8. When the majority of cells appear to have detached, quickly add an equal volume of the Trypsin Neutralizing Solution (ATCC PCS-999-004) to each flask. Gently pipette or swirl the culture to ensure all of the trypsin-EDTA solution has been neutralized.
9. Transfer the dissociated cells to a sterile centrifuge tube and set aside while processing any remaining cells in the culture flask.
10. Add 3 to 5 mL D-PBS (ATCC 30-2200) to the tissue culture flask to collect any additional cells that might have been left behind.
11. Transfer the cell/D-PBS suspension to the centrifuge tube containing the trypsin-EDTA-dissociated cells.
12. Repeat steps 10 and 11 as needed until all cells have been collected from the flask.
13. Centrifuge the cells at 150 x g for 3 to 5 minutes.
14. Aspirate neutralized dissociation solution from the cell pellet and resuspend the cells in 2 to 8 mL fresh, pre-warmed, complete growth medium.
15. Count the cells and seed new culture flasks at a density of 5,000 viable cells per cm2. 16. Place newly seeded flasks in a 37°C, 5% CO2 incubator for at least 24 to 48 hours before processing the cells further. Refer to Maintenance>/i> for guidelines on feeding.
Volume 1.0 mL
Cells per Vial One vial contains a minimum of 5 x 105 cells
Sterility Tests Bacteria and Yeast: Negative
Mycoplasma: Negative
Viral Testing Hepatitis B: Negative
Hepatitis C: Negative
HIV: Negative
Population Doubling Time Refer to Certificate of Analysis for lot specific data
C of A
Certificate of Analysis
Certificate of Analysis
Notice: Necessary PermitsPermits

These permits may be required for shipping this product:

  • Customers located in the state of Hawaii will need to contact the Hawaii Department of Agriculture to determine if an Import Permit is required. A copy of the permit or documentation that a permit is not required must be sent to ATCC in advance of shipment.
Basic Documentation
Product Sheet
Product Sheet
Certificate of Analysis
Certificate of Analysis
SDS
SDS