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Code: 0111
Cell Line: HT-29
Species: Homo sapiens
Tissue: Colon
Morphology: Epithelial
Growth Properties: Adherent
Applications: This cell line is a suitable transfection host.
Products: secretory component of IgA; carcinoembryonic antigen (CEA); transforming growth factor beta binding protein; mucin
Biosafety: 1
Additional info: Ultrastructural features reported for HT-29 cells include microvilli, microfilaments, large vacuolated mitochondria with dark granules, smooth and rough endoplasmic reticulum with free ribosomes, lipid droplets, few primary and many secondary lysosomes. The cells express urokinase receptors, but do not have detectable plasminogen activator activity [PubMed ID: 8381394]. HT-29 cells are negative for CD4, but there is cell surface expression of galactose ceramide (a possible alternative receptor for HIV). The line is positive for expression of c-myc, K-ras, H-ras, N-ras, Myb, sis and fos oncogenes. The p53 antigen is overproduced, and there is a G -> A mutation in codon 273 of the p53 gene resulting in an Arg -> His substitution. N-myc oncogene expression was not detected. There is a G -> A mutation in codon 273 of the p53 gene resulting in an Arg -> His substitution.
Culture Medium: McCoy's 5A Medium is modified to contain 1.5 mM L-glutamine and 2200 mg/L sodium bicarbonate and fetal bovine serum to a final concentration of 10%.
Subculturing: Remove and discard culture medium. Briefly rinse the cell layer with PBS without calcium and magnesium to remove all traces of serum that contains trypsin inhibitor. Add 2.0 to 3.0 mL of Trypsin-EDTA solution to flask and observe cells under an inverted microscope until cell layer is dispersed (usually within 5 to 15 minutes). Note: To avoid clumping do not agitate the cells by hitting or shaking the flask while waiting for the cells to detach. Cells that are difficult to detach may be placed at 37°C to facilitate dispersal. Add 6.0 to 8.0 mL of complete growth medium and aspirate cells by gently pipetting. Add appropriate aliquots of the cell suspension to new culture vessels. Incubate cultures at 37°C. 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.   
Culture Conditions: Atmosphere: air, 95%; carbon dioxide (CO2), 5% Temperature: 37°C
Cryopreservation: 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).
References: Didier ES, et al. Characterization of Encephalitozoon (Septata) intestinailis isolates cultured from nasal mucosa and bronchoalveolar lavage fluids of two AIDS patients. J. Eukaryot. Microbiol. 43: 34-43, 1996. PubMed: 8563708 Fogh J. Human tumor cells in vitro. New York: Plenum Press; 1975. Chen TR, et al. WiDr is a derivative of another colon adenocarcinoma cell line, HT-29. Cancer Genet. Cytogenet. 27: 125-134, 1987. PubMed: 3472642 Fogh J, et al. Absence of HeLa cell contamination in 169 cell lines derived from human tumors. J. Natl. Cancer Inst. 58: 209-214, 1977. PubMed: 833871 Goodfellow M, et al. One hundred and twenty-seven cultured human tumor cell lines producing tumors in nude mice. J. Natl. Cancer Inst. 59: 221-226, 1977. PubMed: 77210034 Adachi A, et al. Productive, persistent infection of human colorectal cell lines with human immunodeficiency virus. J. Virol. 61: 209-213, 1987. PubMed: 3640832 Fantini J, et al. Human colon epithelial cells productively infected with human immunodeficiency virus show impaired differentiation and altered secretion. J. Virol. 66: 580-585, 1992. PubMed: 1727501 Butzow R, et al. A 60-kD protein mediates the binding of transforming growth factor-beta to cell surface and extracellular matrix proteoglycans. J. Cell Biol. 122: 721-727, 1993. PubMed: 8335695 Trainer DL, et al. Biological characterization and oncogene expression in human colorectal carcinoma cell lines. Int. J. Cancer 41: 287-296, 1988. PubMed: 3338874 Hanski C, et al. Tumorigenicity, mucin production and AM-3 epitope expression in clones selected from the HT-29 colon carcinoma cell line. Int. J. Cancer 50: 924-929, 1992. PubMed: 1372882 Reiter LS, et al. The role of the urokinase receptor in extracellular matrix degradation by HT29 human colon carcinoma cells. Int. J. Cancer 53: 444-450, 1993. PubMed: 8381394 Barnett SW, et al. Characterization of human immunodeficiency virus type 1 strains recovered from the bowel of infected individuals. Virology 182: 802-809, 1991. PubMed: 2024498 Shabahang M, et al. 1,25-Dihydroxyvitamin D3 receptor as a marker of human colon carcinoma cell line differentiation and growth inhibition. Cancer Res. 53: 3712-3718, 1993. PubMed: 8393379 Lesuffleur T, et al. Differential expression of the human mucin genes MUC1 to MUC5 in relation to growth and differentiation of different mucus-secreting HT- 29 cell subpopulations. J. Cell Sci. 106: 771-778, 1993. PubMed: 8308060 Pollack MS, et al. HLA-A, B, C and DR alloantigen expression on forty-six cultured human tumor cell lines. J. Natl. Cancer Inst. 66: 1003-1012, 1981. PubMed: 7017212 Fantini J, et al. Infection of colonic epithelial cell lines by type 1 human immunodeficiency virus is associated with cell surface expression of galactosylceramide, a potential alternative gp120 receptor. Proc. Natl. Acad. Sci. USA 90: 2700-2704, 1993. PubMed: 8464878 Devedjian JC, et al. Regulation of the alpha 2A-adrenergic receptor in the HT29 cell line. Effects of insulin and growth factors. J. Biol. Chem. 266: 14359-14366, 1991. PubMed: 1677644 Santoro IM, Groden J. Alternative splicing of the APC gene and its association with terminal differentiation. Cancer Res. 57: 488-494, 1997. PubMed: 9012479 Bermudez LE, et al. Exposure to low oxygen tension and increased osmolarity enhance the ability of Mycobacterium avium to enter intestinal epithelial (HT-29) cells. Infect. Immun. 65: 3768-3773, 1997. PubMed: 9284150 Tsao H, et al. Novel mutations in the p16/CDKN2A binding region of the Cyclin-dependent Kinase-4 gene. Cancer Res. 58: 109-113, 1998. PubMed: 9426066 Qian XC, Brent TP. Methylation hot spots in the 5' flanking region denote silencing of the O6-methylguanine-DNA methyltransferase gene. Cancer Res. 57: 3672-3677, 1997. PubMed: 9288770 Morin PJ, et al. Apoptosis and APC in colorectal tumorigenesis. Proc. Natl. Acad. Sci. USA 93: 7950-7954, 1996. PubMed: 8755583 White LJ, et al. Attachment and entry of recombinant norwalk virus capsids to cultured human and animal cell lines. J. Virol. 70: 6589-6597, 1996. PubMed: 8794293 Kolanus W, et al. alphaLbeta2 integrin/LFA-1 binding to ICAM-1 induced by cytohesin-1 a cytoplasmic regulatory molecule. Cell 86: 233-242, 1996. PubMed: 8706128 Wang R, et al. Cellular adherence elicits ligand-independent activation of the Met cell-surface receptor. Proc. Natl. Acad. Sci. USA 93: 8425-8430, 1996. PubMed: 8710887 Young SW, et al. Gadolinium(III) texaphyrin: a tumor selective radiation sensitizer that is detectable by MRI. Proc. Natl. Acad. Sci. USA 93: 6610-6615, 1996. PubMed: 8692865 Groh V, et al. Cell stress-regulated human major histocompatibility complex class I gene expressed in gastrointestinal epithelium. Proc. Natl. Acad. Sci. USA 93: 12445-12450, 1996. PubMed: 8901601 Takahashi K, et al. Keratan sulfate modification of CD44 modulates adhesion to hyaluronate. J. Biol. Chem. 271: 9490-9496, 1996. PubMed: 8621620
Depositors: LÚCIO MENDES CABRAL - Unviersidade Federal do Rio de Janeiro