hct-116、hela、MEFs细胞参数介绍.doc

HELAHeLa (ATCCCCL-2)ATCC：Hela细胞，源于黑人31岁女性，子宫颈腺癌，是一种附着型上皮细胞，要求存于液氮中。These cells are a suitable transfection host.This cell line can be used to screen forEscherichia colistrains with invasive potential.Biosafety Level：（2 Cells contain human papilloma virusBiosafety 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.）Complete Growth Medium：The base medium for this cell line is ATCC-formulated Eagles Minimum Essential Medium, Catalog No. 30-2003. To make the complete growth medium, add the following components to the base medium: fetal bovine serum to a final concentration of 10%.Subculturing（接种）：Volumes used in this protocol are for a 75 cm2flask; proportionally reduce or increase amount of dissociation medium for culture vessels of other sizes. CorningT-75 flasks (catalog #430641) are recommended for subculturing this product.1. Remove and discard culture medium.2. Briefly rinse the cell layer with 0.25% (w/v) Trypsin- 0.53 mM EDTA solution to remove all traces of serum which contains trypsin inhibitor.3. 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 37C to facilitate dispersal.4. Add 6.0 to 8.0 mL of complete growth medium and aspirate cells by gently pipetting.5. Add appropriate aliquots of the cell suspension to new culture vessels.6. Incubate cultures at 37C.Subcultivation Ratio:A subcultivation ratio of 1:2 to 1:6 is recommendedMedium Renewal:2 to 3 times per weekCryopreservationFreeze Medium:Complete growth medium supplemented with 5% (v/v) DMSOStorage Temperature:Liquid nitrogen vapor phaseCulture ConditionsAtmosphere:Air, 95%; carbon dioxide (CO2), 5%Temperature:37C论坛：贴壁生长，铺路石状，长得快，2-3天传代一次，传代不及时会造成老化的细胞堆积，看起来很脏。10%+1640或者10%+高糖DMEM都有培养。MEFs_283TAg (ATCCCRL-2822)OrganismMus musculus, mouseTissueembryoCell Typefibroblast immortalized with SV40 large T antigenSV40 large T antigen transfectedProduct FormatfrozenMorphologyfibroblastCulture PropertiesadherentBiosafety Level2 cells containing SV40 viral DNA sequencesBiosafety classification is based on聽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.Age14.5 day gestation embryoApplicationsDNA repair studiesStorage Conditionsliquid nitrogen vapor phaseDerivation283TAg (Polb/Mpg double null) is a mouse embryonic fibroblast (MEF) cell line derived from polymerase (DNA directed), beta (Polb)/ N-methylpurine-DNA glycosylase (Mpg, Aag) double null embryos at day 14.5 of gestation. The cell line was established by transfection with an expression vector for SV40 large T antigen PubMed: 8538772. The cells are transgenic for lambda LIZ (Lac I/cII).Comments283TAg (Polb/Mpg double null) is a mouse embryonic fibroblast (MEF) cell line derived from polymerase (DNA directed), beta (Polb)/ N-methylpurine-DNA glycosylase (Mpg, Aag) double null embryos at day 14.5 of gestation. The cell line was established by transfection with an expression vector for SV40 large T antigen PubMed: 8538772. The cells are transgenic for lambda LIZ (Lac I/cII).Complete Growth MediumThe base medium for this cell line is ATCC-formulated Dulbeccos Modified Eagles Medium, Catalog No. 30-2002. To make the complete growth medium, add the following components to the base medium: fetal bovine serum to a final concentration of 10%.SubculturingProtocol:1. Remove and discard culture medium.2. Briefly rinse the cell layer with 0.25% (w/v) Trypsin- 0.53 mM EDTA solution to remove all traces of serum that contains trypsin inhibitor.3. 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 37C to facilitate dispersal.4. Add 6.0 to 8.0 ml of complete growth medium and aspirate cells by gently pipetting.5. Add appropriate aliquots of the cell suspension to new culture vessels.An inoculum of 4 X 10(3) to 4 X 10(4) viable cells/cm2 is recommended.6. Incubate cultures at 37C.Interval:Maintain cultures at a cell concentration between 6 X 10(3) and 1 X 10(5) cells/cm2.Subcultivation Ratio:A subcultivation ratio of 1:6 to 1:8 is recommendedMedium Renewal:Two to three times weeklyCryopreservationFreeze medium:Complete growth medium supplemented with 5% (v/v) DMSOStorage temperature:liquid nitrogen vapor phaseCulture ConditionsAtmosphere:air, 95%; carbon dioxide (CO2), 5%Temperature:37.0CPopulation Doubling Time29 hoursHCT 116 (ATCCCCL-247)OrganismHomo sapiens, humanTissuecolonProduct FormatfrozenMorphologyepithelialCulture PropertiesadherentBiosafety Level1Biosafety classification is based on聽U.S. Public Health Service Guidelines,聽it is the responsibility of the customer to ensure that their facilities comply with biosafety regulations for their own country.Diseasecolorectal carcinoma(结肠直肠癌)AgeadultGendermaleApplicationsThis cell line is a suitable transfection host.This line has a mutation in codon 13 of the ras proto-oncogene, and can be used as a positive control for PCR assays of mutation in this codon.Storage Conditionsliquid nitrogen vapor phaseKaryotypeThe stemline chromosome number is near diploid with the modal number at 45 (62%) and polyploids occurring at 6.8%. The markers 10q+ and t(?8p;18q) are present in all metaphases and t(9q;?16p-), in 80% of the cells karyotyped. N16 is monosomic in the presence of, but disomic in the absence of t(9q;?16p-). N10 and N18 are monosomic and other chromosomes from those mentioned above are disomic. Q-band observations revealed the presence of the Y chromosome, but not in all cells (50% of cells lacked the Y in G-band karyotypes).ImagesClinical DatamaleGenes Expressedcarcinoembryonic antigen (CEA) 1 ng per 106聽cells per 10 days.Cellular Productscarcinoembryonic antigen (CEA) 1 ng per 10 exp6 cells per 10 days; keratinTumorigenicYesEffectsYes, in nude miceRefCommentsThe cells are positive for keratin by immunoperoxidase staining.HCT 116 cells are positive for transforming growth factor beta 1 (TGF beta 1) and beta 2 (TGF beta 2) expression.Complete Growth MediumThe base medium for this cell line is ATCC-formulated McCoys 5a Medium Modified, Catalog No. 30-2007. To make the complete growth medium, add the following components to the base medium: fetal bovine serum to a final concentration of 10%.聽SubculturingVolumes are given for a 75 cm2聽flask. Corning庐聽T-75 flasks (catalog #430641) are recommended for subculturing this product. Increase or decrease the amount of dissociation medium needed proportionally for culture vessels of other sizes.1. Remove and discard culture medium.2. Briefly rinse the cell layer with 0.25% (w/v) Trypsin- 0.53 mM EDTA solution to remove all traces of serum which contains trypsin inhibitor.3. 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 d