MDCK-SIAT1

MDCK-SIAT1 were permanently transfected with the gene of the human CMP-N-acetylneuraminate beta-galactoside-2,6-sialyltransferase (SIAT1) (EC 2.4.99.1), an enzyme that catalyzes the alfa-2,6-sialylation of N-acetyllactosamine moieties of glycoproteins. As a result, clinical human and swine influenza viruses bind better to MDCK-SIAT1 cells than to MDCK cells.

Strain: Cocker spaniel 

Virus Susceptibility: Human and swine influenza viruses

The base medium for this cell line is ATCC-formulated DNEN Medium (ATCC 30-2002). To make the complete growth medium, add the following components to the base medium:

• Fetal bovine (ATCC 30-2020) serum to a final concentration of 10%.
• L-glutamine (ATCC 30-2214) to a final concentration of 2mM
• GibcoTM G-418 (ThermoFisher catalog # 10131-027) to a final concentration of 1 mg/mL

Note: Because of the limited stability of G-418, we recommend spiking in fresh G-418 prior to seeding or performing fluid additions. Media supplemented with G-418 should not be stored. Complete culture medium is stable for 7 days at 37°C.

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 culture medium without G-418 and spin at approximately 150 to 400 x g for 8 to 12 minutes.
4. Carefully aspirate the supernatant and discard, leaving the cell pellet.
5. Gently resuspend the cell pellet with the appropriate amount of growth medium without G-418 and transfer cell suspension into a vented T-75.
6. Place the flask in a 37°C incubator with 5% CO₂ .
7. On day 1 on culture, replace the media with complete culture medium containing G-418)

1. Remove and discard culture medium.
2. Briefly rinse the cell layer with PBS to remove all traces of serum that contains trypsin inhibitor.
3. Add 2.0 to 3.0 mL of Trypsin-EDTA for Primary Cells (ATCC PCS-999-003) to flask and observe cells under an inverted microscope until the cell layer is dispersed (usually up to 30 minutes).
   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 aspirate cells by gently pipetting.
5. Centrifuge the cell suspension at approximately 150 to 400 x g for 8 to 12 minutes to remove dissociation agent.
6. Resuspend the cell pellet in an appropriate amount of complete culture medium
7. Add appropriate aliquots of the cell suspension to new culture vessels.
   Cultures can be established between 1.0 x 10⁴ and 4.0 x 10⁴ viable cells/cm².
8. Incubate cultures at 37°C.

The product is provided 'AS IS' and the viability of ATCC^® products is warranted for 30 days from the date of shipment, provided that the customer has stored and handled the product according to the information included on the product information sheet, website, and Certificate of Analysis. For living cultures, ATCC lists the media formulation and reagents that have been found to be effective for the product. While other unspecified media and reagents may also produce satisfactory results, a change in the ATCC and/or depositor-recommended protocols may affect the recovery, growth, and/or function of the product. If an alternative medium formulation or reagent is used, the ATCC warranty for viability is no longer valid.  Except as expressly set forth herein, no other warranties of any kind are provided, express or implied, including, but not limited to, any implied warranties of merchantability, fitness for a particular purpose, manufacture according to cGMP standards, typicality, safety, accuracy, and/or noninfringement.

This product is intended for laboratory research use only. It is not intended for any animal or human therapeutic use, any human or animal consumption, or any diagnostic use. Any proposed commercial use is prohibited without a license from ATCC.

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