CuFi-1細胞

運輸方式： 凍存運輸

細胞形態： 上皮樣

ATCC Number： CRL-4013?

相關**： 囊腫性纖維化

數量： 大量

細胞類型： 上皮細胞

是否是腫瘤細胞： 0

物種來源： 人

生長狀態： 貼壁生長

年限： 14

器官來源： 肺（及支氣管）

Designations: CuFi-1

Depositors: AJ Klingelhutz

CuFi-1細胞Biosafety Level: 2 [cells containing SV40 and HPV viral DNA sequences ]

Shipped: frozen

Medium & Serum: See Propagation

Growth Properties: adherent

Organism: Homo sapiens

Morphology: epithelial

Source: Organ: lung; bronchus

Disease: cystic fibrosis

Cell Type: epithelial

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Isolation: Isolation date: 2000

Applications: These cell lines may be useful models for studying ion physiology, therapeutic interventions for cystic fibrosis and innate immunity [Pubmed: 12676769].

CuFi-1細胞As part of our quality control, we have tested this cell line for its ability to grow for a minimum of 15 population doublings after recovery from cryopreservation.

We have also compared its karyotype, telomerase expression level, growth rate, morphology and tissue-specific markers when first recovered from cryopreservation with that of cells at 10+ population doublings to ensure that there is no change in these parameters and that the cells are capable of extended proliferation.

Both cell lines, when seeded on semipermeable filters and grown at the air-liquid interface, are capable of forming polarized differentiated epithelia that exhibit transepithelial resistance and maintain the ion channel physiology expected of each genotype.

Human airway epithelial (HAE) cell line, CuFi-1, was derived from lung of a 14-year-old patient with cystic fibrosis by dual retroviral infection with HPV-16E6/E7-LXSN [Pubmed: 1845902] and hTERT-LXSN [Pubmed: 9817205].

DNA Profile (STR): Amelogenin: X

CSF1PO: 12

D13S317: 11,13

D16S539: 11,14

D5S818: 12

D7S820: 8,11

THO1: 6,9.3

TPOX: 8,11

vWA: 17,20

Cytogenetic Analysis: CuFi-1細胞This is a near-diploid human cell line of female origin with a modal chromosome count of 46 and a polyploidy rate of 27%. There were two copies of a karyotypically normal X-chromosome present in most of the cells. Overall, some of the cells contained chromosomal abnormalities, with most consistent being trisomy 20.

Age: 14

Gender: female

Comments: Human airway epithelial (HAE) cell line, CuFi-1, was derived from lung of a 14-year-old patient with cystic fibrosis by dual retroviral infection with HPV-16E6/E7-LXSN [Pubmed: 1845902] and hTERT-LXSN [Pubmed: 9817205]. Consequently, the cells do not undergo growth arrest in cell culture due to exogenous expression of the telomerase and E6/E7 genes. CuFi-1 cells are homozygous for the delta F508 cystic fibrosis-causing mutation (delta F508/delta F508). Another hTERT-immortalized cell line, derived from normal HAE is also available as ATCC CRL-4011 (NuLi-1). Both cell lines, when seeded on semipermeable filters and grown at the air-liquid interface, are capable of forming polarized differentiated epithelia that exhibit transepithelial resistance and maintain the ion channel physiology expected of each genotype. These cell lines may be useful models for studying ion physiology, therapeutic interventions for cystic fibrosis and innate immunity [Pubmed: 12676769].

As part of our quality control, we have tested this cell line for its ability to grow for a minimum of 15 population doublings after recovery from cryopreservation. We have also compared its karyotype, telomerase expression level, growth rate, morphology and tissue-specific markers when first recovered from cryopreservation with that of cells at 10+ population doublings to ensure that there is no change in these parameters and that the cells are capable of extended proliferation.

Propagation: ATCC complete growth medium: These cells are grown in a serum-free medium: BEGM (Bronchial Epithelial Growth Medium, Serum-free) from Lonza (BEGM Bullet Kit; CC-3170) made of BEBM basal medium and SingleQuot additives (ATCC does not use gentamycin-amphotericin B) supplemented with 50 ?g/ml G-418.

Atmosphere: 5% CO2 in air recommended

Temperature: 37.0°C

Subculturing: Protocol:

Note: CuFi-1細胞The culture flasks should be pre-coated with 60ug/ml solution of Human Placental Collagen Type IV. (Sigma Cat. No. C-7521) at least 18 hours in advance then air-dried and rinsed 2-3 times with Dulbecco?s Phosphate Buffered Saline.

Volumes used in this protocol are for 75 sq.cm. flasks; proportionally reduce or increase amount of dissociation medium for culture vessels of other sizes.

Remove and discard culture medium.

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.

To remove Trypsin-EDTA solution, add 2.0 to 3.0 ml of 1% FBS in Dulbecco's Phosphate buffered Saline and aspirate cells by gently pipetting.

Transfer cell suspension to a centrifuge tube and spin at approximately 125 x g for 5 to 10 minutes. Discard supernatant.

Resuspend the cell pellet in fresh growth medium. Add appropriate aliquots of the cell suspension to new culture vessels. An inoculum of 1 x 10(3) to 3 x 10(3) viable cells/sq. cm is recommended.

Incubate cultures at 37C.

Subculture when cell concentration is between 1 x 10(4) and 2 x 10(4) cells/sq. cm.

Subcultivation Ratio: 1:5

Medium Renewal: Every 2-3 days (do not exceed 3 days)

Preservation: Freeze medium: BEGM with 30% FBS and 10% DMSO

Storage temperature: liquid nitrogen vapor phase

Doubling Time: about 26 hours

Related Products: recommended serum:ATCC 30-2020

0.25% (w/v) Trypsin - 0.53 mM EDTA in Hank' BSS (w/o Ca++, Mg++):ATCC 30-2101

phosphate-buffered saline:CuFi-1細胞ATCC 30-2200

Cell culture tested DMSO:ATCC 4-X

Erythrosin B vital stain solution:ATCC 30-2404

References: 22566: Halbert CL, et al. The E7 gene of human papillomavirus type 16 is sufficient for immortalization of human epithelial cells. J. Virol. 65: 473-478, 1991. PubMed: 1845902

47354: Bodnar AG, et al. Extension of life-span by introduction of telomerase into normal human cells. Science 279: 349-352, 1998. PubMed: 9454332

92666: Zabner J, et al. Development of cystic fibrosis and noncystic fibrosis airway cell lines . Am. J. Physiol. Lung Cell Mol Physiol. 284: L844-L854, 2003. PubMed: 12676769

92667: Kiyono T, et al. Both Rb/p161NK4a inactivation and telomerase activity are required to immortalize human epithelial cells. Nature 396: 84-88, 1998. PubMed: 9817205