1、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. Publ
2、ic 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 t
3、he 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. Cor
4、ningT-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 sol
5、ution 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
6、 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:
7、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
8、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 classifi
9、cation 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 phaseDerivation
10、283TAg (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
11、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
12、 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.
13、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 tra
14、ces 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 wai
15、ting 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
16、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 supple
