Ninety-six-well 0.45 μm Hydrophobic Multiscreen plates (Milipore) were coated with αIFN-γ mAb (clone 1-D1K, Mabtech) overnight at 4°C. The plate was blocked for 2 h after which 20 000 C1R cells transfected with CD1a or CD1b were co-cultured with 200 T cells in the present or absence of a blocking antibody (clone BCD1b.3) and different antigens. The antigens used in this assay are BbGL-II lipid (5 μg/mL), sonicated B. burgdorferi (200 000 bacteria per well) and media as control. After incubation overnight at 37°C cells were lysed and washed away with PBS-Tween, and the plates were incubated for 2 h with a biotinylated αIFN-γ antibody (clone 7-B6–1, Mabtech). The wells were washed with PBS-Tween and incubated with Extravidin-ALP (Sigma–Aldrich) for 1 h. After washing with PBS-Tween followed by washing with PBS, 5-Bromo-4-chloro-3-indolyl phosphate/Nitro blue tetrazolium (SigmaFAST tablets, Sigma) was added to visualize the spots. Spots were counted by the Immunospot reader (C.T.L Technologies).
αifn γ mab clone 1 d1k
Manufactured by Mabtech
The αIFN-γ mAb (clone 1-D1K) is a monoclonal antibody that binds to interferon-gamma (IFN-γ). This antibody can be used in laboratory applications for the detection and quantification of IFN-γ.
Automatically generated - may contain errors
2 protocols using αifn γ mab clone 1 d1k
1
ELISPOT Assay for CD1-Restricted T Cells
2
IFN-γ ELISPOT Assay for CD1-Restricted T Cells
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Ninety‐six‐well 0.45 μm Hydrophobic Multiscreen plates (Milipore) were coated with αIFN‐γ mAb (clone 1‐D1K, Mabtech) overnight at 4°C. The plate was blocked for 2 h after which 20 000 C1R cells transfected with CD1a or CD1b were co‐cultured with 200 T cells in the present or absence of a blocking antibody (clone BCD1b.3) and different antigens. The antigens used in this assay are BbGL‐II lipid (5 μg/mL), sonicated B. burgdorferi (200 000 bacteria per well) and media as control. After incubation overnight at 37°C cells were lysed and washed away with PBS‐Tween, and the plates were incubated for 2 h with a biotinylated αIFN‐γ antibody (clone 7‐B6‐1, Mabtech). The wells were washed with PBS‐Tween and incubated with Extravidin‐ALP (Sigma–Aldrich) for 1 h. After washing with PBS‐Tween followed by washing with PBS, 5‐Bromo‐4‐chloro‐3‐indolyl phosphate/Nitro blue tetrazolium (SigmaFAST tablets, Sigma) was added to visualize the spots. Spots were counted by the Immunospot reader (C.T.L Technologies).
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