||Cervix, Hela Contaminant
||Originally derived from an epidermoid carcinoma of the mouth of an adult male Caucasian. It was one of the early attempts to isolate and serially propagate a human cell line directly on glass as a monolayer. NB: This cell line contains HeLa marker chromosomes and expresses type A G6PD. For further information see Nature 1976;259:211; In Vitro 1978;14:469.
||The cells are positive for keratin by immunoperoxidase staining.
KB cells have been reported to contain human papillomavirus 18 (HPV-18) sequences.
NOTE: Cells of this line contain HeLa marker chromosomes, and were derived via HeLa contamination
||Dulbecco's Modified Eagle's Medium (DMEM) with 1% non-essential amino acids, 2 mM L-glutamine, 1.0 g/L glucose and 10% of fetal bovine serum.
||Remove medium, and rinse with PBS without calcium and magnesium.
Remove the solution and add an additional 1 to 2 mL of trypsin-EDTA solution. Allow the flask to sit at room temperature (or at 37°C) until the cells detach.
Add fresh culture medium, aspirate and dispense into new culture flasks.
NOTE: For more information on enzymatic dissociation and subculturing of cell lines consult Chapter 12 in Culture of Animal Cells, a manual of Basic Technique by R. Ian Freshney, 6th edition, published by Alan R. Liss, N.Y., 2010.
|Subculturing Medium Renewal
||2 to 3 times per week
|Subculturing Subcultivation Ratio
||1:4 to 1:10
||Atmosphere: air, 95%; carbon dioxide (CO2), 5%
||95% FBS + 5% DMSO (Dimethyl sulfoxide)
|Thawing Frozen Cells
||SAFETY PRECAUTION: Is highly recommend that protective gloves and clothing always be used and a full face mask always be worn when handling frozen vials. It is important to note that
some vials leak when submersed in liquid nitrogen and will slowly fill with liquid nitrogen. Upon thawing, the conversion of the liquid nitrogen back to its gas phase may result in the vessel exploding or blowing off its cap with dangerous force creating flying debris.
1. Thaw the vial by gentle agitation in a 37°C water bath. To reduce the possibility of contamination, keep the Oring and cap out of the water. Thawing should be rapid (approximately 2 minutes).
2. Remove the vial from the water bath as soon as the contents are thawed, and decontaminate by dipping in or spraying with 70% ethanol. All of the operations from this point on should be carried out under strict aseptic conditions.
3. For cells that are sensitive to DMSO is recommended that the cryoprotective agent be removed immediately. Transfer the vial contents to a centrifuge tube containing 9.0 mL complete culture medium and spin at approximately 125 x g for 5 to 7 minutes.
4.Discard the supernatant and Resuspend cell pellet with the recommended complete medium (see the specific batch information for the culture recommended dilution ratio).
5. Incubate the culture in a appropriate atmosphere and temperature (see "Culture Conditions" for this cell line).
NOTE: It is important to avoid excessive alkalinity of the medium during recovery of the cells. It is suggested that, prior to the addition of the vial contents, the culture vessel containing the growth medium be placed into the incubator for at least 15 minutes to allow the medium to reach its normal pH (7.0 to 7.6).
||Eagle H. Propagation in a fluid medium of a human epidermoid carcinoma, strain KB. Proc. Soc. Exp. Biol. Med. 89: 362-364, 1955. PubMed: 13254761
Boshart M, et al. A new type of papillomavirus DNA, its presence in genital cancer biopsies and in cell lines derived from cervical cancer. EMBO J. 3: 1151-1157, 1984. PubMed: 6329740
Salzman NP. Animal Cell Cultures. Science 133:1559-65. 1961 PubMed: 13745868
Eagle H, Foley GE. Cytotoxicity in human cell cultures as a primary screen for the detection of anti-tumor agents. Cancer Res. 18: 1017-1025, 1958. PubMed: 13596943
Foley GE, Handler AH. Differentiation of “Normal” and Neoplastic Cells Maintained in Tissue Culture by Implantation into Normal Hamsters. Proc Soc Exp Biol Med 94: 661-664 1957. PubMed: 13431913
Eagle H, et al. Viral susceptibility of a human carcinoma cell (strain KB). Proc. Soc. Exp. Biol. Med. 91: 361-364, 1956. PubMed: 13322936
St. Geme JW, et al. Characterization of the genetic locus encoding Haemophilus influenzae type b surface fibrils. J. Bacteriol. 178: 6281-6287, 1996. PubMed: 8892830
Chang K, Pastan I. Molecular cloning of mesothelin, a differentiation antigen present on mesothelium, mesotheliomas, and ovarian cancers. Proc. Natl. Acad. Sci. USA 93: 136-140, 1996. PubMed: 8552591
Lee RJ, Huang L. Folate-targeted, anionic liposome-entrapped polylysine-condensed DNA for tumor cell-specific gene transfer. J. Biol. Chem. 271: 8481-8487, 1996. PubMed: 8626549
||Jose Paulo Leite, Instituto Oswaldo Cruz, Rio de Janeiro.