品系： Landrace

ATCC Number： CRL-2843?

細胞形態： 單核細胞/巨噬細胞

生長狀態： 貼壁生長

年限： 27 days

運輸方式： 凍存運輸

數量： 大量

器官來源： 肺

細胞類型： 其他細胞類型

是否是腫瘤細胞： 0

物種來源： 豬

Designations: 3D4/21

3D4/21細胞Depositors: J Gren

Biosafety Level: 2 [cells containing SV40 viral DNA sequences ]

Shipped: frozen

Medium & Serum: See Propagation

Growth Properties: adherent

Organism: Sus scrofa

Morphology: macrophage

Source: Organ: lung

Strain: Landrace

Cell Type: macrophage macrophage (alveolar); immortalized with SV40 large T antigentransformed with pSV3-neo

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Isolation: Isolation date: December, 1998

Applications: These porcine myelomonocytic cell lines may have a wide variety of applications in porcine virology and immunology [PubMed:12088830].

3D4/21細胞Clone 3D4/21 can produce Bovine adenovirus type 3 (BAV-3) to markedly higher titers than clones 3D4/2 and 3D4/31. Addition of DMSO improved the capability of clone 3D4/21 to replicate the field isolate of African swine fever virus (ASFV/Lillie) compared to the other clones.

Single cell cloning and selection in G-418 of the 3D4 parental cell line resulted in establishment of 3D4/2 (ATCC CRL-2845), 3D4/21 (ATCC CRL-2843) and 3D4/31 (ATCC CRL-2844).

The parental porcine monomyeloid cell line, 3D4, was established in December of 1998 following transfection of primary porcine alveolar macrophage cultures with the pSV3neo plasmid.

A subpopulation of each cell line was positive, to varying degrees depending on the media formulation, for nonspecific esterase activity and phagocytosis.

Virus Susceptibility: Bovine adenovirus 3

Classical swine fever virus

Human parainfluenza virus 3

Swinepox virus

Vesicular stomatitis New Jersey virus

Porcine adenovirus

Herpes simplex virus 1

African swine fever virus

Pseudorabies virus

Vaccinia virus

Swine vesicular disease virus

Age: 27 days

Gender: unknown

Comments: 3D4/21細胞The parental porcine monomyeloid cell line, 3D4, was established in December of 1998 following transfection of primary porcine alveolar macrophage cultures with the pSV3neo plasmid. The plasmid carries the genes for neomycin resistance and SV40 large T antigen. Single cell cloning and selection in G-418 of the 3D4 parental cell line resulted in establishment of 3D4/2 (ATCC CRL-2845), 3D4/21 (ATCC CRL-2843) and 3D4/31 (ATCC CRL-2844). A subpopulation of each cell line was positive, to varying degrees depending on the media formulation, for nonspecific esterase activity and phagocytosis. Clone 3D4/21 can produce Bovine adenovirus type 3 (BAV-3) to markedly higher titers than clones 3D4/2 and 3D4/31. Addition of DMSO improved the capability of clone 3D4/21 to replicate the field isolate of African swine fever virus (ASFV/Lillie) compared to the other clones. These porcine myelomonocytic cell lines may have a wide variety of applications in porcine virology and immunology [PubMed:12088830].

Propagation: ATCC complete growth medium: RPMI 1640 medium with 2 mM L-glutamine adjusted to contain 1.5 g/L sodium bicarbonate, 4.5 g/L glucose, 10 mM HEPES, 1.0 mM sodium pyruvate supplemented with 0.1 mM nonessential amino acids, 90%; fetal bovine serum, 10%

Atmosphere: 5% CO2 in air recommended

Temperature: 37.0°C

Subculturing: Protocol: Volumes used in this protocol are for 75cm2 flasks; proportionally reduce or increase amount of dissociation medium for culture vessels of other sizes.

Remove and discard culture medium.

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.

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.3D4/21細胞 An inoculum of 5 X 10(3) to 7 X 10(3) viable cells/cm2 is recommended.

Incubate cultures at 37?C. Subculture when cell concentration reaches between 3 X 10(5) and 4 X 10(5) cells/cm2.

Subcultivation Ratio: A subcultivation ratio of 1:6 to 1:8 is recommended

Medium Renewal: Two to three times weekly

Preservation: Freeze medium: Complete growth medium supplemented with 5% (v/v) DMSO

Storage temperature: liquid nitrogen vapor phase

Doubling Time: about 18 hrs

Related Products: Recommended medium (without the additional supplements or serum described under ATCC Medium):ATCC 30-2001

recommended serum:ATCC 30-2020

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

Cell culture tested DMSO:ATCC 4-X

Erythrosin B vital stain solution:ATCC 30-2404

Trypan Blue vital stain solution:ATCC 30-2402

MEM Non-Essential Amino Acid Solution, 100x:ATCC 30-2116

References: 92770: Weingartl HM, et al. 3D4/21細胞Continuous porcine cell lines developed from alveolar macrophages: partial characterization and virus susceptibility. J. Virol. Methods 104: 203-216, 2002. PubMed: 12088830