Normal Human CD8+ Cytotoxic T Cells INTRODUCTION
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1. fuge for 5 min at 400 x g Aspirate or decant the supernatant and gently resuspend the cell pellet in 10 ml of medium Remove an aliquot for cell count and proceed with experimental manipulations Culture the cells in RPMI 1640 medium supplemented with 10 v v FBS 2 mM glutamine 1 v v nonessential amino acids 1 v v sodium pyruvate 50 U ml penicillin 50 mg ml streptomycin and 50 mg ml kanamycin at a density of 1 2 million cells ml NOTE The cell suspension may form clumps after standing at room temperature This can be avoided by preparing and using the cells promptly or by adding DNase to the suspension at a final concentration of 10 units per ml Normal Human CD8 Cytotoxic T Cells Manual C References 1 Inflammatory disease protective R381Q IL23 receptor polymorphism results in decreased primary CD4 and CD8 human T cell functional responses Ritu Sarin Xingxin Wu and Clara Abraham PNAS June 7 201 Vol 108 No 23 9560 9565 VKM120202 Genlantis Page 2 of 2 10190 Telesis Court San Diego CA 92121 Toll Free 888 428 0558 U S amp Canada 858 457 1919 www genlantis com 2011 Genlantis and Gene Therapy Systems Inc2. ids 1 v v sodium pyruvate 50 U ml penicillin 50 mg ml streptomycin and 50 mg ml kanamycin B Thawing and Culturing Cells 1 2 3 Prepare a 37 C water bath to temperature Keep all samples frozen until the bath is ready Place the vials into the water bath being careful not to submerge below the junction between the lid and the vial Place vial of cells in 37 C water bath and agitate until thawed It is important to thaw the cells quickly do NOT allow thawed cells to remain in freezing media any longer than necessary When only a small amount of ice remains remove the vials and dry with a lab tissue Clean the top of the vial with a lab tissue wetted with 70 alcohol avoid wiping away the labeling Within about 30 sec slowly add one milliliter of medium containing serum to the thawed cells antis l division of Gene Therapy Systems Inc Normal Human Peripheral Blood Mononuclear PBMC000 5 0x10 Cells PBMC001 25 0x10 PBMC002 75 0x108 Normal Human CD4 Helper T Cells PBMC003A 10 0x106 Normal Human CD14 Monocytes PBMC005A 10 0x108 Normal Human CD19 B Cells PBMCOO6A 10 0x108 Normal Human CD56 Natural Killer Cells Normal Human Dendritic Cells PBMC007A 10 0x106 PBMCO008A 0 5x106 Slowly add thawed cells to 8 ml of medium containing serum Invert tube 2 or 3 times to mix or mix gently by pipetting up and down several times Centri
3. mmunoPure HD lt 4 PBMCO004A Normal Human CD8 Cytotoxic T 10 0 x 108 cells Cells negatively selected Cryopreserved Cells are shipped on dry ice For maximum viability it is best to use cells as soon as possible For short term 3 weeks or less store cells at 80 C for longer term store cells in liquid nitrogen 170 C NOTE viability cannot be guaranteed following 170 C storage or beyond one week at 80 C INTRODUCTION Normal Human CD8 Cytotoxic T Cells are a sub group of T lymphocytes capable of inducing the death of infected with viruses or other pathogens somatic or tumor cells Most CD8 Cytotoxic T cells express T cell receptors that can recognize a specific antigenic peptide bound to Class MHC molecules present on all nucleated cells and a glycoprotein called CD8 The MHC molecule keeps the Cytotoxic T cell and the target cell bound closely together during antigen specific activation CD8 T cells are recognized as Cytotoxic T cells once they become activated and are generally classified as having a pre defined cytotoxic role within the immune system The ImmunoPure HD CD8 Cytotoxic T Cells are isolated from fresh human adult peripheral blood of healthy donors at IRB approved and FDA licensed blood banks using apheresis and immunomagnetic separation techniques MATERIALS AND METHODS A General Medium Requirement RPMI 1640 medium 10 v v FBS 2 mM glutamine 1 v v nonessential amino ac
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