X-ray-irradiated DC (2 × 104) and the T cells (1 × 105) were seeded into wells of 96-well flat-bottom culture plates in RPMI 1640 10% fetal bovine serum (Invitrogen Life Technologies) for 3 days. Supernatants were removed and further analyzed for cytokine production (IL-2 and IFN-γ) with ELISA. Plates were read at 405 nm using a Labsystems Multiskan enzyme-linked immunosorbent assay reader.
Rpmi 1640 10 fetal bovine serum
Manufactured by Thermo Fisher Scientific
RPMI 1640 10% fetal bovine serum is a cell culture medium that provides essential nutrients for the growth and maintenance of cells in vitro. It contains RPMI 1640 base medium supplemented with 10% fetal bovine serum, which supplies growth factors and other components to support cell proliferation.
Automatically generated - may contain errors
4 protocols using rpmi 1640 10 fetal bovine serum
1
Cytokine Production in DC-T Cell Co-culture
2
Characterizing T Cell Activation in Mixed Lymphocyte Reaction
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Cells activated in an MLR were allowed to incubate for up to 3 days before harvesting.
[3H]Thymidine(0.5μCi/well; MPBiomedicals) was added 24h before harvesting (SkatronInstruments) using Type A lter mats (Perkin-Elmer Life and Analytical Sciences) and a beta plate scintillation mixture (Perkin-Elmer). Disintegrations per minute were determined using a liquid scintillation counter (1205 Betaplate; Perkin-Elmer).
ELISA assay for quantitating cytokine production X-ray-irradiated DC (2×10 4 ) and the T cells (1×10 5 ) were seeded into wells of 96-well at-bottom culture plates in RPMI 1640 10% fetal bovine serum (Invitrogen Life Technologies) for 3 days. Supernatants were removed and further analyzed for cytokine production (IL-2 and IFN-γ) with ELISA. Plates were read at 405 nm using a Labsystems Multiskan enzyme-linked immunosorbent assay reader.
Flow cytometry for detecting activation marker expression FCM assay was used to characterize activation marker expression of T cells. CD4 + or CD8 + T cells were cocultured with DCs for 3 days. The cells were washed with PBS and then stained with anti-CD3-PE, anti-CD25-FITC and anti-CD69-APC at 4 •C for 1h, and then washed by PBS. CD25 and CD69 activation markers were measured using a FCM Calibur ow cytometer (BD company). Data were analyzed with CellQuest software.
[3H]Thymidine(0.5μCi/well; MPBiomedicals) was added 24h before harvesting (SkatronInstruments) using Type A lter mats (Perkin-Elmer Life and Analytical Sciences) and a beta plate scintillation mixture (Perkin-Elmer). Disintegrations per minute were determined using a liquid scintillation counter (1205 Betaplate; Perkin-Elmer).
ELISA assay for quantitating cytokine production X-ray-irradiated DC (2×10 4 ) and the T cells (1×10 5 ) were seeded into wells of 96-well at-bottom culture plates in RPMI 1640 10% fetal bovine serum (Invitrogen Life Technologies) for 3 days. Supernatants were removed and further analyzed for cytokine production (IL-2 and IFN-γ) with ELISA. Plates were read at 405 nm using a Labsystems Multiskan enzyme-linked immunosorbent assay reader.
Flow cytometry for detecting activation marker expression FCM assay was used to characterize activation marker expression of T cells. CD4 + or CD8 + T cells were cocultured with DCs for 3 days. The cells were washed with PBS and then stained with anti-CD3-PE, anti-CD25-FITC and anti-CD69-APC at 4 •C for 1h, and then washed by PBS. CD25 and CD69 activation markers were measured using a FCM Calibur ow cytometer (BD company). Data were analyzed with CellQuest software.
3
Comprehensive Immune Response Profiling
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Cells activated in an MLR were allowed to incubate for up to 3 days before harvesting.
[3H]Thymidine(0.5 µCi/well; MPBiomedicals) was added 24 h before harvesting (SkatronInstruments) using Type A lter mats (Perkin-Elmer Life and Analytical Sciences) and a beta plate scintillation mixture (Perkin-Elmer). Disintegrations per minute were determined using a liquid scintillation counter (1205 Betaplate; Perkin-Elmer).
ELISA assay for quantitating cytokine production X-ray-irradiated DC (2 × 10 4 ) and the T cells (1 × 10 5 ) were seeded into wells of 96-well at-bottom culture plates in RPMI 1640 10% fetal bovine serum (Invitrogen Life Technologies) for 3 days. Supernatants were removed and further analyzed for cytokine production (IL-2 and IFN-γ) with ELISA. Plates were read at 405 nm using a Labsystems Multiskan enzyme-linked immunosorbent assay reader.
Flow cytometry for detecting activation marker expression FCM assay was used to characterize activation marker expression of T cells. CD4 + or CD8 + T cells were cocultured with DCs for 3 days. The cells were washed with PBS and then stained with anti-CD3-PE, anti-CD25-FITC and anti-CD69-APC at 4 •C for 1 h, and then washed by PBS. CD25 and CD69 activation markers were measured using a FCM Calibur ow cytometer (BD company). Data were analyzed with CellQuest software.
[3H]Thymidine(0.5 µCi/well; MPBiomedicals) was added 24 h before harvesting (SkatronInstruments) using Type A lter mats (Perkin-Elmer Life and Analytical Sciences) and a beta plate scintillation mixture (Perkin-Elmer). Disintegrations per minute were determined using a liquid scintillation counter (1205 Betaplate; Perkin-Elmer).
ELISA assay for quantitating cytokine production X-ray-irradiated DC (2 × 10 4 ) and the T cells (1 × 10 5 ) were seeded into wells of 96-well at-bottom culture plates in RPMI 1640 10% fetal bovine serum (Invitrogen Life Technologies) for 3 days. Supernatants were removed and further analyzed for cytokine production (IL-2 and IFN-γ) with ELISA. Plates were read at 405 nm using a Labsystems Multiskan enzyme-linked immunosorbent assay reader.
Flow cytometry for detecting activation marker expression FCM assay was used to characterize activation marker expression of T cells. CD4 + or CD8 + T cells were cocultured with DCs for 3 days. The cells were washed with PBS and then stained with anti-CD3-PE, anti-CD25-FITC and anti-CD69-APC at 4 •C for 1 h, and then washed by PBS. CD25 and CD69 activation markers were measured using a FCM Calibur ow cytometer (BD company). Data were analyzed with CellQuest software.
4
Mixed Lymphocyte Reaction and Cytokine Quantification
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Cells activated in an MLR were allowed to incubate for up to 3 days before harvesting.
[3H]Thymidine(0.5μCi/well; MPBiomedicals) was added 24h before harvesting (SkatronInstruments) using Type A lter mats (Perkin-Elmer Life and Analytical Sciences) and a beta plate scintillation mixture (Perkin-Elmer). Disintegrations per minute were determined using a liquid scintillation counter (1205 Betaplate; Perkin-Elmer).
ELISA assay for quantitating cytokine production X-ray-irradiated DC (2×10 4 ) and the T cells (1×10 5 ) were seeded into wells of 96-well at-bottom culture plates in RPMI 1640 10% fetal bovine serum (Invitrogen Life Technologies) for 3 days. Supernatants were removed and further analyzed for cytokine production (IL-2 and IFN-γ) with ELISA. Plates were read at 405 nm using a Labsystems Multiskan enzyme-linked immunosorbent assay reader.
[3H]Thymidine(0.5μCi/well; MPBiomedicals) was added 24h before harvesting (SkatronInstruments) using Type A lter mats (Perkin-Elmer Life and Analytical Sciences) and a beta plate scintillation mixture (Perkin-Elmer). Disintegrations per minute were determined using a liquid scintillation counter (1205 Betaplate; Perkin-Elmer).
ELISA assay for quantitating cytokine production X-ray-irradiated DC (2×10 4 ) and the T cells (1×10 5 ) were seeded into wells of 96-well at-bottom culture plates in RPMI 1640 10% fetal bovine serum (Invitrogen Life Technologies) for 3 days. Supernatants were removed and further analyzed for cytokine production (IL-2 and IFN-γ) with ELISA. Plates were read at 405 nm using a Labsystems Multiskan enzyme-linked immunosorbent assay reader.
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