For Th17 cell generation, cells were stimulated by plate–bound anti-CD3 (2C11, 5 μg/ml; eBioscience) plus CD28 (37.51, 1 μg/ml; eBioscience), recombinant murine IL6 (rIL6, 20 ng/ml; Peprotech, Rocky Hill, NJ) and recombinant TGF-β (rTGF-β, 0.5 ng/ml; Peprotech) in the presence or absence of vitamin C (10 μg/ml, Sigma-Aldrich, St. Louis, MO) for 3 d. To prevent the Foxp3 expression, 10 μg/ml of anti-IL2 blocking mAb (clone: S4B6) was always added except some samples in supplementary Fig. 1 .
Recombinant tgf β rtgf β
Manufactured by Thermo Fisher Scientific
Sourced in Germany
Recombinant TGF‐β (rTGF‐β) is a laboratory reagent produced using recombinant DNA technology. It is a member of the transforming growth factor beta (TGF‐β) family of proteins. TGF‐β proteins play a role in various cellular processes, including cell growth, differentiation, and immune function.
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Lab products found in correlation
Anti type 1 collagen af 5610 antibody, by OriGene (3 mentions)
Dimethylmethylene blue dye, by Serva Electrophoresis (2 mentions)
Abc reagent, by Vector Laboratories (2 mentions)
Anti type x collagen col 10 antibody, by Merck Group (2 mentions)
Anti cd3 2c11, by Thermo Fisher Scientific (1 mentions)
Vitamin c, by Merck Group (1 mentions)
Biotinylated secondary antibody, by Vector Laboratories (1 mentions)
Htgf β quantikine elisa, by R&D Systems (1 mentions)
Anti type x collagen col 10, by Merck Group (1 mentions)
Anti tgf β 5 and anti sox9 c 20 antibodies, by Santa Cruz Biotechnology (1 mentions)
Fgf 2 quantikine elisa dfb50, by R&D Systems (1 mentions)
Diaminobenzidine dab, by Merck Group (1 mentions)
Anti igf 1 af 291 na antibody, by R&D Systems (1 mentions)
Higf 1 quantikine elisa, by R&D Systems (1 mentions)
Dimethylmethylene blue dmmb, by Serva Electrophoresis (1 mentions)
Anti sox9 c 20 antibody, by Santa Cruz Biotechnology (1 mentions)
Abc kit, by Vector Laboratories (1 mentions)
4 protocols using recombinant tgf β rtgf β
1
In Vitro Generation of Th17 Cells
2
Chondrocyte Extracellular Matrix Analysis
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All reagents were from Sigma‐Aldrich (Munich, Germany), unless otherwise identified. Recombinant TGF‐β (rTGF‐β) was purchased at Peprotech (Hamburg, Germany) and the dimethylmethylene blue dye at Serva (Heidelberg, Germany). The anti‐TGF‐β (V) and anti‐SOX9 (C‐20) antibodies were from Santa Cruz Biotechnology (Heidelberg, Germany), the anti‐type‐II collagen (II‐II6B3) antibody from the NIH Hybridoma Bank (University of Iowa, Ames, USA), the anti‐type‐I collagen (AF‐5610) antibody from Acris (Hiddenhausen, Germany) and the anti‐type‐X collagen (COL‐10) from Sigma‐Aldrich. Biotinylated secondary antibodies and the ABC reagent were obtained from Vector Laboratories (Alexis Deutschland GmbH, Grünberg, Germany). The hTGF‐β Quantikine ELISA was from R&D Systems (Wiesbaden, Germany) and the type‐II, ‐I and ‐X collagen ELISAs from from Antibodies‐Online (Aachen, Germany).
3
Chondrogenic Differentiation Assay
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All reagents were from Sigma (Munich, Germany), unless otherwise identified. Recombinant TGF-β (rTGF-β) was purchased at Peprotech (Hamburg, Germany) and the dimethylmethylene blue dye at Serva (Heidelberg, Germany). The anti-FGF-2 (C-18) and anti-SOX9 (C-20) antibodies were from Santa Cruz Biotechnology (Heidelberg, Germany), the anti-type-II collagen (II-II6B3) antibody from the NIH Hybridoma Bank (University of Iowa, Ames, USA), the anti-type-I collagen (AF-5610) antibody from Acris (Hiddenhausen, Germany), and the anti-type-X collagen (COL-10) antibody from Sigma. Biotinylated secondary antibodies and the ABC reagent were obtained from Vector Laboratories (Alexis Deutschland GmbH, Grünberg, Germany). The FGF-2 Quantikine ELISA (DFB50) was from R&D Systems (Wiesbaden, Germany) and the type-II, −I, and -X collagen ELISAs from Antibodies-Online (Aachen, Germany).
4
Extracellular Matrix Characterization Protocol
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Recombinant TGF‐β (rTGF‐β) was purchased at Peprotech (Hamburg, Germany), dimethylmethyleneblue (DMMB) at Serva (Heidelberg, Germany) and diaminobenzidine (DAB) at Sigma‐Aldrich (Munich, Germany). The anti‐IGF‐I (AF‐291‐NA) antibody was from R&D Systems (Wiesbaden, Germany), the anti‐type‐II collagen (II‐II6B3) antibody from NIH Hybridoma Bank (University of Iowa, Ames, USA), the anti‐SOX9 (C‐20) antibody from Santa Cruz Biotechnology (Heidelberg, Germany), the anti‐type‐I collagen (AF‐5610) antibody from Acris (Hiddenhausen, Germany) and the anti‐type‐X collagen (COL‐10) antibody from Sigma‐Aldrich. Secondary biotinylated antibodies and the ABC kit were from Vector Laboratories (Alexis Deutschland GmbH, Grünberg, Germany). The hIGF‐I Quantikine ELISA was purchased at R&D Systems and the type‐II, ‐I and ‐X collagen ELISAs at Antibodies‐Online (Aachen, Germany).
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