黄色小说视频-日本少妇高潮抽搐-国内自拍av-天堂中文资源在线-蜜桃视频在线入口www-91久久综合-亚洲视频在线一区-日日夜夜拍-国产精品资源在线观看-欧美激情国产精品免费-人妻少妇久久中文字幕-成人免费无码大片a毛片-香蕉污视频在线观看-日本电影成人-国内精品久久99人妻无码

The Foreign Animal Disease Diagnostic Laboratory of the US Department of Agriculture has determined that ATCC CCL-33 is not infected with Hog cholera virus or African swine fever virus, and uses this line to screen for those viruses.

The cell line harbors an endogenous C-type retrovirus.

The cells are positive for porcine circovirus (PCV) antigens.

The cells are positive for keratin by immunoperoxidase staining.

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

1. Remove and discard culture medium.
2. Briefly rinse the cell layer twice with 0.25% (w/v) Trypsin-0.53mM 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 37°C 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 37°C.

Subcultivation Ratio: A subcultivation ratio of 1:2 to 1:4 is recommended

Dulac GC, Afshar A. Porcine circovirus antigens in PK-15 cell line (ATCC CCL-33) and evidence of antibodies to circovirus in Canadian pigs. Can. J. Vet. Res. 53: 431-433, 1989. PubMed: 2686830

Pirtle EC, Woods LK. Cytogenetic alterations in swine kidney cells persistently infected with hog cholera virus and propagated with and without antiserum in the medium. Am. J. Vet. Res. 29: 153-164, 1968. PubMed: 4965860

Armstrong JA, et al. C-type virus particles in pig kidney cell lines. J. Gen. Virol. 10: 195-198, 1971. PubMed: 4324256

Newman JT, Smith KO. Characteristics of a swine papovavirus. Infect. Immun. 5: 961-967, 1972. PubMed: 4344097

Tumilowicz JJ, et al. Concurrent replication of a papovavirus and a C-type virus in the CCL 33 porcine cell line. In Vitro 15: 922-928, 1979. PubMed: 232060

Todaro GJ, et al. Characterization of a type C virus released from the porcine cell line PK(15). Virology 58: 65-74, 1974. PubMed: 4132403

James A House, personal communication

Nucleotide (GenBank) : AJ133817 Porcine endogenous retrovirus type C proviral gag, pol and env genesand LTR (class A, clone 42).

Nucleotide (GenBank) : AJ133816 Porcine endogenous retrovirus type C proviral gag, pol and env genes and LTR (class B, clone 33).