首页 » HelaRC32 cell line重组腺相关病毒包装细胞株[HeRC32]-BioVector NTCC质粒载体菌株细胞蛋白抗体基因保藏中心CRL-2972

HelaRC32 cell line重组腺相关病毒包装细胞株[HeRC32]-BioVector NTCC质粒载体菌株细胞蛋白抗体基因保藏中心CRL-2972

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HelaRC32 cell line重组腺相关病毒包装细胞株[HeRC32]-BioVector NTCC质粒载体菌株细胞蛋白抗体基因保藏中心


HeLaRC32 [HeRC32] is an epithelial-like cell that was isolated from the cervix of a Black, 31-year-old patient with adenocarcinoma. This cell line was deposited by P Moullier.


Product category
Human cells
Organism
Homo sapiens, human
Morphology
epithelial
Tissue
Uterus; Cervix
Disease
Adenocarcinoma
Applications
3D cell culture
Product format
Frozen
Storage conditions
Vapor phase of liquid nitrogen


General

Specific applications

These cells generate rAAV preparations with high titers of infectious particles which are essentially free of adenoviruses for biological, preclinical, clinical or pharmaceutical applications.

HelaRC32 is a stable packaging cell line expressing the rep and cap genes for recombinant adeno-associated virus type 2 (rAAV-2) assembly which constitutes an attractive alternative to transient transfection protocols.

Characteristics

Growth properties
Adherent
Derivation
This is a clone of HeLa cells that harbors one to two rep-cap gene copies per cell.
Age
31 years
Ethnicity
Black
Gender
Female
Genes expressed
Rep proteins: (Rep 78; Rep 68; Rep 52; and Rep 40); Cap proteins: (VP1; VP2; and VP3)
Comments

This is a clone of HeLa cells that harbors one to two rep-cap gene copies per cell. Upon vector transfection and adenovirus infection, efficient rAAV assembly correlated with a 100-fold amplification of the integrated rep-cap sequence with the inverted terminal repeats (ITRs) deleted.


Handling information

Unpacking and storage instructions
  1. Check all containers for leakage or breakage.

  2. Remove the frozen cells from the dry ice packaging and immediately place the cells at a temperature below ­-130°C, preferably in liquid nitrogen vapor, until ready for use.

Complete medium
The base medium for this cell line is ATCC-formulated Dulbecco's Modified Eagle's 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%.
Handling procedure

To insure the highest level of viability, thaw the vial and initiate the culture as soon as possible upon receipt. If upon arrival, continued storage of the frozen culture is necessary, it should be stored in liquid nitrogen vapor phase and not at -70°C.  Storage at -70°C will result in loss of viability.

  1. Thaw the vial by gentle agitation in a 37°C water bath. To reduce the possibility of contamination, keep the O-ring and cap out of the water. Thawing should be rapid (approximately 2 minutes).

  2. Remove the vial from the water bath as soon as the contents are thawed, and decontaminate by dipping in or spraying with 70% ethanol. All of the operations from this point on should be carried out under strict aseptic conditions.

  3. Transfer the vial contents to a centrifuge tube containing 9.0 mL complete culture medium and spin at approximately 125 x g for 5 to 10 minutes.

  4. Resuspend cell pellet with the recommended complete medium (see the specific batch information for the culture recommended dilution ratio) and dispense into a 25 cm2 or a 75 cm2 culture flask.  It is important to avoid excessive alkalinity of the medium during recovery of the cells. It is suggested that, prior to the addition of the vial contents, the culture vessel containing the complete growth medium be placed into the incubator for at least 15 minutes to allow the medium to reach its normal pH (7.0 to 7.6).

  5. Incubate the culture at 37°C in a suitable incubator.  A 5% CO2 in air atmosphere is recommended if using the medium described on this product.

Subculturing procedure
Volumes used in this protocol are for 75 cm2 flasks; 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 with Ca++/Mg++ free Dulbecco's phosphate-buffered saline (D-PBS) or 0.25% (w/v) Trypsin - 0.53 mM EDTA solution to remove all traces of serum which contains trypsin inhibitor.

  3. Add 1.0 to 2.0 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. Transfer cell suspension to a centrifuge tube and spin at approximately 125 X g for 5 to 10 minutes. Discard supernatant.

  6. Resuspend the cell pellet in fresh growth medium. Add appropriate aliquots of the cell suspension to new culture vessels.

  7. Incubate cultures at 37°C.

Subcultivation ratio: A subcultivation ratio of 1:2 to 1:6 is recommended.
Medium renewal: Every 2 to 3 days
Reagents for cryopreservation
Complete culture medium + an additional 10% fetal bovine serum (FBS) + 5% DMSO

Quality control specifications

Mycoplasma contamination
Not detected
STR profiling
Amelogenin: X
CSF1PO: 9,10
D13S317: 12,13.3
D16S539: 9,10
D5S818: 11,12
D7S820: 8,12
TH01: 7
TPOX: 8,12
vWA: 16,18
D2S1338: 17
D19S433: 13,14
FGA: 21
D8S1179: 12,13
Penta_D: 8,15
Penta_E: 7,17
D18S51: 16
D21S11: 27,28
D3S1358: 15,18

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