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Product Name:
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LuSIV Cell Line
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Manufacturer:
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Virongy Biosciences, Inc., Manassas, Virginia, USA
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Material Provided:
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Each vial contains approximately 1.0 mL of frozen cells in 90% fetal bovine serum and 10% dimethylsulfoxide (DMSO). Sufficient cells are provided to initiate at least one new culture. The cell count, expressed as cells/vial, is shown on individual Certificates of Analysis for each lot.
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Packing/Storage:
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This product was packaged aseptically in cryovials. It should be stored at -100°C or colder, preferably in the vapor phase of a liquid nitrogen freezer. Storage at -70°C will result in loss of viability. To ensure the highest level of viability, the vial should be thawed and the culture initiated as soon as possible upon receipt. Any warming of the product during shipping and transfer must be avoided, as this will adversely affect the viability of the product after thawing. For the transfer between freezers and shipping, the cells may be placed on dry ice for brief periods, although the use of a portable liquid nitrogen carrier is preferred. Please read the following recommendations prior to reconstituting this material.
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Safety Precautions:
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When handling frozen vials, it is highly recommended that protective gloves, lab coat and full-face mask be worn. Even brief exposure to the ultra-cold temperature can cause tissue damage from frostbite. Also, some vials may slowly fill with liquid nitrogen if they have been immersed during cryogenic storage. When thawing, the liquid nitrogen may rapidly expand as it changes to gas, breaking the vial or cap with explosive force, sending debris flying with enough velocity to cause injury. Store and use in areas with adequate ventilation
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Thawing and Growth:
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Note: This is a suspension cell line. Do not discard floating cells.
Prior to thawing the cells, prepare growth medium (GM) for use. HRP-5460 is typically grown in RPMI-1640 medium supplemented with 10% FBS, 2 mM sodium pyruvate, 10 mM HEPES, 2 mM L-glutamine, 0.5 mg/mL gentamicin and 300 µg/mL hygromycin B. Omit the hygromycin B for the first 24 hours of growth.
Rapidly thaw the vial of cells in a 37°C water bath with gentle agitation. To reduce the risk of contamination, keep the cap and O-ring of the vial out of the water and repeatedly check the cap for tightness during thawing. Remove from the water bath immediately when thawed. Dry the vial with a sterile wiper, decontaminate using a wiper soaked with 70% isopropyl alcohol, and let the vial air dry. Aseptically open the vial, remove the vial contents and add to 4.0 mL of GM in a centrifuge tube. Centrifuge the cell suspension at 150 × g for 8 to 10 minutes at 18°C to 25°C. Discard the supernatant and resuspend the cell pellet in 10 mL of pre-warmed GM without hygromycin B. Transfer the cell suspension into a 75 cm2 tissue culture flask. Incubate the new culture at 37°C and 5% CO2.
Allow LuSIV cells to grow without hygromycin B for 24 hours. Split cells 1:5 every three to four days. The cells are a suspension cell line that grows in clumps. Propagation media should contain 300 µg/mL hygromycin B for maintenance of reporter plasmid. For optimal sensitivity, LuSIV cells should not be passaged more than 25 times.
After 24 hours, perform a total media change by placing the flask contents into a centrifuge tube. Centrifuge the cell suspension at 150 × g for 8 to 10 minutes at 18°C to 25°C. Discard the supernatant and resuspend the cell pellet in 10 mL of pre-warmed GM with hygromycin B.
Sub-culture procedure.
Cultures can be maintained by the addition of fresh GM or the replacement of GM. Alternatively, cultures can be established by centrifugation with subsequent resuspension at 1.0 × 105 cells/mL. Maintain a cell density between 1.0 × 105 and 1.0 × 106 cells/mL. Centrifuge the cell suspension at 150 × g for 8 to 10 minutes at 18°C to 25°C. Discard the supernatant and resuspend the cell pellet in 10 mL of pre-warmed GM to the desired cell density. Add fresh medium every 2 to 3 days, depending on cell density.
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Disclaimers:
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You are authorized to use this product for research use only. It is not intended for human use. Use of this product is subject to the terms and conditions of the BEI Resources Material Transfer Agreement (MTA). The MTA is available on our Web site at www.beiresources.org. While BEI Resources uses reasonable efforts to include accurate and up-to-date information on this product sheet, neither ATCC® nor the U.S. Government makes any warranties or representations as to its accuracy. Citations from scientific literature and patents are provided for informational purposes only. Neither ATCC® nor the U.S. Government warrants that such information has been confirmed to be accurate. This product is sent with the condition that you are responsible for its safe storage, handling, use and disposal. ATCC® and the U.S. Government are not liable for any damages or injuries arising from receipt and/ or use of this product. While reasonable effort is made to ensure authenticity and reliability of materials on deposit, the U.S. Government, ATCC®, their suppliers and contributors to BEI Resources are not liable for damages arising from the misidentification or misrepresentation of products. |
References:
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1. Roos, J. W., et al. “LuSIV Cells: A Reporter Cell Line for the Detection and Quantitation of a Single Cycle of HIV and SIV Replication.” Virology 273 (2000): 307-315. PubMed: 10915601.
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Citation:
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Acknowledgment for publications should read “The following reagent was obtained through BEI Resources, NIAID, NIH: LuSIV Cell Line, HRP-5460."
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Biosafety Level:
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1
Appropriate safety procedures should always be used with this material. Laboratory safety is discussed in the following publication: U.S. Department of Health and Human Services, Public Health Service, Centers for Disease Control and Prevention, and National Institutes of Health. Biosafety in Microbiological and Biomedical Laboratories (BMBL). Current Edition. Washington, DC: U.S. Government Printing Office.
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