Normal Human CD4+ Helper T Cells INTRODUCTION MATERIALS
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1. PBMC003A Normal Human CD4 Helper T Cells 10 0 x 10 cells 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 3s ImmunoPure HD lt 4 A division of Gene Therapy Systems Inc Normal Human Peripheral Blood Mononuclear PBMC000 5 0x10 Cells PBMC001 25 0x10 PBMC002 75 0x108 Normal Human CD8 Cytotoxic T cells PBMCOO4A 10 0x108 Normal Human CD14 Monocytes PBMCO05A 10 0x108 Normal Human CD19 B Cells PBMCOO6A 10 0x108 Normal Human CD56 Natural Killer Cells PBMC007A 10 0x108 Normal Human Dendritic Cells PBMCO08A 0 5x108 Normal Human CD4 Helper T cells are a sub group of lymphocytes that play a crucial role in establishing and maximizing the capabilities of the immune system These cells are unusual in that they have no cytotoxic or phagocytic activity they cannot kill infected host cells also known as somatic cells or pathogens CD4 T cells are essential in determining B cell antibody class switching in the activation and growth of cytotoxic T cells and in maximizing bactericidal activity of phagocytes such as macrophages It is this diversity in function and their role in influ2. encing other cells that gives T helper cells their name The ImmunoPure HD Human CD4 Helper 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 cell separation techniques MATERIALS AND METHODS A General Medium Requirement RPMI 1640 medium 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 B Thawing and Culturing Cells 1 Prepare a 37 C water bath to temperature 2 Keep all samples frozen until the bath is ready 3 Place the vials into the water bath being careful not to submerge below the junction between the lid and the vial 4 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 5 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 moistened with 70 alcohol avoid wiping away the labeling 6 Within about 30 sec slowly add one milliliter of medium containing serum to the thawed cells 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 Centrifuge for 5 min at 400 x g Aspirate or decant the supe
3. rnatant 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 CD4 Helper T Cells Manual C References 1 CXCR5 Expressing Human Central Memory CD4 T Cells and Their Relevance for Humoral Immune Responses Nina Chevalier David Jarrossay Edwin Ho Danielle T Avery Cindy S Ma Di Yu Federica Sallusto Stuart G Tangye and Charles R Mackay J Immunol 2011 186 5556 5568 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 Inc
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