Flow Cytometry:Article Title: An engineered linear cap-independent mRNA vaccine with intrinsic adjuvanticity induces potent anti-tumor immunity in mice.
Article Snippet: .. Flow cytometry analysis AR TI CL E IN P RE SS For OVA (SIINFEKL) Tetramer staining, PBMCs were separated by Mouse 1× Lymphocyte Separation Medium (DAKEWE, Shenzhen) according to the manufacturer’s instructions. .. Cells were then stained with Fixable Viability Dye eFluor 450 (65-0863-14, eBioscience), anti-CD3 (APC-65077, Proteintech), anti-CD8a (FITC65069, Proteintech), H-2Kb&B2M&OVA (SIINFEKL) Tetramer (PE, H2A-MP2H7-25ug, ACROBiosystems), and analyzed by CytoFLEX Flow Cytometer (Beckman).
Staining:Article Title: An engineered linear cap-independent mRNA vaccine with intrinsic adjuvanticity induces potent anti-tumor immunity in mice.
Article Snippet: .. Flow cytometry analysis AR TI CL E IN P RE SS For OVA (SIINFEKL) Tetramer staining, PBMCs were separated by Mouse 1× Lymphocyte Separation Medium (DAKEWE, Shenzhen) according to the manufacturer’s instructions. .. Cells were then stained with Fixable Viability Dye eFluor 450 (65-0863-14, eBioscience), anti-CD3 (APC-65077, Proteintech), anti-CD8a (FITC65069, Proteintech), H-2Kb&B2M&OVA (SIINFEKL) Tetramer (PE, H2A-MP2H7-25ug, ACROBiosystems), and analyzed by CytoFLEX Flow Cytometer (Beckman).
Article Title: An engineered linear cap-independent mRNA vaccine with intrinsic adjuvanticity induces potent anti-tumor immunity in mice.
Article Snippet: Cells were then stained with Fixable Viability Dye eFluor 450 (65-0863-14, eBioscience), anti-CD3 (APC-65077, Proteintech), anti-CD8a (FITC65069, Proteintech), anti-mouse CD4 (PE-CY7, 116016 , Biolegend), H-2Kb&B2M&OVA (SIINFEKL) Tetramer (PE, H2A-MP2H7-25ug, ACROBiosystems) , and analyzed by CytoFLEX Flow Cytometer (Beckman). .. For intra-cellular IFNγ staining, PBMCs or splenocytes were separated by Mouse 1× Lymphocyte Separation Medium (DAKEWE, Shenzhen) according to the manufacturer’s instructions. .. Cells were then stained with Fixable Viability Dye eFluor 450 (65-0863-14, eBioscience), anti-CD3 (APC-65077, Proteintech), anti-CD8a at 4°C for 30min.
Article Title: An engineered linear cap-independent mRNA vaccine with intrinsic adjuvanticity induces potent anti-tumor immunity in mice
Article Snippet: .. For OVA (SIINFEKL) Tetramer staining, PBMCs were separated by Mouse 1× Lymphocyte Separation Medium (DAKEWE, Shenzhen) according to the manufacturer’s instructions. .. Cells were then stained with Fixable Viability Dye eFluor 450 (65-0863-14, eBioscience), anti-CD3 (APC-65077, Proteintech), anti-CD8a (FITC-65069, Proteintech), H-2Kb&B2M&OVA (SIINFEKL) Tetramer (PE, H2A-MP2H7-25ug, ACROBiosystems), and analyzed by CytoFLEX Flow Cytometer (Beckman).
Article Title: Engineering Linear Cap-independent mRNA Vaccines with Intrinsic Adjuvanticity for Potent Cancer Immunotherapy
Article Snippet: .. For OVA (SIINFEKL) Tetramer staining, PBMCs was separated by Mouse 1× Lymphocyte Separation Medium (DAKEWE, Shenzhen) according to the manufacturer’s instruction. .. Cells were then stained with Fixable Viability Dye eFluor 450 (65-0863-14, eBioscience), anti-CD3 (APC-65077, Proteintech), anti-CD8a (FITC-65069, Proteintech), H-2Kb&B2M&OVA (SIINFEKL) Tetramer (PE, ACROBiosystems), and analyzed by CytoFLEX Flow Cytometer (Beckman).
Article Title: Enhanced Th1 Cellular Immunity Induced by an RSV-F mRNA Vaccine Rationally Designed Using NLP Algorithms.
Article Snippet: .. Lymphocytes were isolated using Mouse 1× Lymphocyte Separation Medium (Dakewe, Cat. #7211011, Dakewe Biotech Co., Ltd., Shenzhen, China), washed twice with PBS, and stimulated with a Respiratory Syncytial Virus Fusion Protein Peptide pool (Vazyme, Cat. #DD9123, 2 μg/mL, Vazyme Biotech Co., Ltd., Nanjing, China) plus CD28 (BD Biosciences, Cat. #553294, BD Biosciences, San Jose, CA, USA) for 18 h. Brefeldin A (BFA) was added and maintained for the final 5 h. First, the cells were stained with a Zombie Aqua Fixable Viability Kit (Biolegend, Cat. #423102, 10 min, RT, BioLegend, Inc., San Diego, CA, USA), followed by APC/Cyanine7 anti-mouse https://doi.org/10.3390/vaccines14040356 CD3 (Biolegend, Cat. #100330), PerCP anti-mouse CD4 (Biolegend, Cat. #100434), and FITC anti-mouse CD8 (Biolegend, Cat. #100706) for 15–30 min at RT. .. After fixation (Biolegend Fixation Buffer, Cat. #420801) and permeabilization (Biolegend Intracellular Staining Permeabilization Wash Buffer, Cat. #421002), intracellular cytokines were stained with PEconjugated anti-mouse IL-2 (Biolegend, Cat. #503808), BV605-conjugated anti-mouse IL-4 (Biolegend, Cat. #504126), APC-conjugated anti-mouse TNF-α (Biolegend, Cat. #506308), and BV421-conjugated anti-mouse IFN-γ (Biolegend, Cat. #505830) for 15 min at RT.
Article Title: Enhanced Th1 Cellular Immunity Induced by an RSV-F mRNA Vaccine Rationally Designed Using NLP Algorithms
Article Snippet: .. Lymphocytes were isolated using Mouse 1× Lymphocyte Separation Medium (Dakewe, Cat. #7211011, Dakewe Biotech Co., Ltd., Shenzhen, China), washed twice with PBS, and stimulated with a Respiratory Syncytial Virus Fusion Protein Peptide pool (Vazyme, Cat. #DD9123, 2 μg/mL, Vazyme Biotech Co., Ltd., Nanjing, China) plus CD28 (BD Biosciences, Cat. #553294, BD Biosciences, San Jose, CA, USA) for 18 h. Brefeldin A (BFA) was added and maintained for the final 5 h. First, the cells were stained with a Zombie Aqua Fixable Viability Kit (Biolegend, Cat. #423102, 10 min, RT, BioLegend, Inc., San Diego, CA, USA), followed by APC/Cyanine7 anti-mouse CD3 (Biolegend, Cat. #100330), PerCP anti-mouse CD4 (Biolegend, Cat. #100434), and FITC anti-mouse CD8 (Biolegend, Cat. #100706) for 15–30 min at RT. .. After fixation (Biolegend Fixation Buffer, Cat. #420801) and permeabilization (Biolegend Intracellular Staining Permeabilization Wash Buffer, Cat. #421002), intracellular cytokines were stained with PE-conjugated anti-mouse IL-2 (Biolegend, Cat. #503808), BV605-conjugated anti-mouse IL-4 (Biolegend, Cat. #504126), APC-conjugated anti-mouse TNF-α (Biolegend, Cat. #506308), and BV421-conjugated anti-mouse IFN-γ (Biolegend, Cat. #505830) for 15 min at RT.
Article Title: An engineered linear cap-independent mRNA vaccine with intrinsic adjuvanticity induces potent anti-tumor immunity in mice
Article Snippet: Cells were then stained with Fixable Viability Dye eFluor 450 (65-0863-14, eBioscience), anti-CD3 (APC-65077, Proteintech), anti-CD8a (FITC-65069, Proteintech), anti-mouse CD4 (PE-CY7, 116016, Biolegend), H-2Kb&B2M&OVA (SIINFEKL) Tetramer (PE, H2A-MP2H7-25ug, ACROBiosystems), and analyzed by CytoFLEX Flow Cytometer (Beckman). .. For intracellular IFN-γ staining, PBMCs or splenocytes were separated by Mouse 1× Lymphocyte Separation Medium (DAKEWE, Shenzhen) according to the manufacturer’s instructions. .. Cells were then stained with Fixable Viability Dye eFluor 450 (65-0863-14, eBioscience), anti-CD3 (APC-65077, Proteintech), and anti-CD8a at 4 °C for 30 min.
Isolation:
Article Title: Enhanced Th1 Cellular Immunity Induced by an RSV-F mRNA Vaccine Rationally Designed Using NLP Algorithms.
Article Snippet: .. Lymphocytes were isolated using Mouse 1× Lymphocyte Separation Medium (Dakewe, Cat. #7211011, Dakewe Biotech Co., Ltd., Shenzhen, China), washed twice with PBS, and stimulated with a Respiratory Syncytial Virus Fusion Protein Peptide pool (Vazyme, Cat. #DD9123, 2 μg/mL, Vazyme Biotech Co., Ltd., Nanjing, China) plus CD28 (BD Biosciences, Cat. #553294, BD Biosciences, San Jose, CA, USA) for 18 h. Brefeldin A (BFA) was added and maintained for the final 5 h. First, the cells were stained with a Zombie Aqua Fixable Viability Kit (Biolegend, Cat. #423102, 10 min, RT, BioLegend, Inc., San Diego, CA, USA), followed by APC/Cyanine7 anti-mouse https://doi.org/10.3390/vaccines14040356 CD3 (Biolegend, Cat. #100330), PerCP anti-mouse CD4 (Biolegend, Cat. #100434), and FITC anti-mouse CD8 (Biolegend, Cat. #100706) for 15–30 min at RT. .. After fixation (Biolegend Fixation Buffer, Cat. #420801) and permeabilization (Biolegend Intracellular Staining Permeabilization Wash Buffer, Cat. #421002), intracellular cytokines were stained with PEconjugated anti-mouse IL-2 (Biolegend, Cat. #503808), BV605-conjugated anti-mouse IL-4 (Biolegend, Cat. #504126), APC-conjugated anti-mouse TNF-α (Biolegend, Cat. #506308), and BV421-conjugated anti-mouse IFN-γ (Biolegend, Cat. #505830) for 15 min at RT.
Article Title: Enhanced Th1 Cellular Immunity Induced by an RSV-F mRNA Vaccine Rationally Designed Using NLP Algorithms
Article Snippet: .. Lymphocytes were isolated using Mouse 1× Lymphocyte Separation Medium (Dakewe, Cat. #7211011, Dakewe Biotech Co., Ltd., Shenzhen, China), washed twice with PBS, and stimulated with a Respiratory Syncytial Virus Fusion Protein Peptide pool (Vazyme, Cat. #DD9123, 2 μg/mL, Vazyme Biotech Co., Ltd., Nanjing, China) plus CD28 (BD Biosciences, Cat. #553294, BD Biosciences, San Jose, CA, USA) for 18 h. Brefeldin A (BFA) was added and maintained for the final 5 h. First, the cells were stained with a Zombie Aqua Fixable Viability Kit (Biolegend, Cat. #423102, 10 min, RT, BioLegend, Inc., San Diego, CA, USA), followed by APC/Cyanine7 anti-mouse CD3 (Biolegend, Cat. #100330), PerCP anti-mouse CD4 (Biolegend, Cat. #100434), and FITC anti-mouse CD8 (Biolegend, Cat. #100706) for 15–30 min at RT. .. After fixation (Biolegend Fixation Buffer, Cat. #420801) and permeabilization (Biolegend Intracellular Staining Permeabilization Wash Buffer, Cat. #421002), intracellular cytokines were stained with PE-conjugated anti-mouse IL-2 (Biolegend, Cat. #503808), BV605-conjugated anti-mouse IL-4 (Biolegend, Cat. #504126), APC-conjugated anti-mouse TNF-α (Biolegend, Cat. #506308), and BV421-conjugated anti-mouse IFN-γ (Biolegend, Cat. #505830) for 15 min at RT.
Virus:Article Title: Enhanced Th1 Cellular Immunity Induced by an RSV-F mRNA Vaccine Rationally Designed Using NLP Algorithms.
Article Snippet: .. Lymphocytes were isolated using Mouse 1× Lymphocyte Separation Medium (Dakewe, Cat. #7211011, Dakewe Biotech Co., Ltd., Shenzhen, China), washed twice with PBS, and stimulated with a Respiratory Syncytial Virus Fusion Protein Peptide pool (Vazyme, Cat. #DD9123, 2 μg/mL, Vazyme Biotech Co., Ltd., Nanjing, China) plus CD28 (BD Biosciences, Cat. #553294, BD Biosciences, San Jose, CA, USA) for 18 h. Brefeldin A (BFA) was added and maintained for the final 5 h. First, the cells were stained with a Zombie Aqua Fixable Viability Kit (Biolegend, Cat. #423102, 10 min, RT, BioLegend, Inc., San Diego, CA, USA), followed by APC/Cyanine7 anti-mouse https://doi.org/10.3390/vaccines14040356 CD3 (Biolegend, Cat. #100330), PerCP anti-mouse CD4 (Biolegend, Cat. #100434), and FITC anti-mouse CD8 (Biolegend, Cat. #100706) for 15–30 min at RT. .. After fixation (Biolegend Fixation Buffer, Cat. #420801) and permeabilization (Biolegend Intracellular Staining Permeabilization Wash Buffer, Cat. #421002), intracellular cytokines were stained with PEconjugated anti-mouse IL-2 (Biolegend, Cat. #503808), BV605-conjugated anti-mouse IL-4 (Biolegend, Cat. #504126), APC-conjugated anti-mouse TNF-α (Biolegend, Cat. #506308), and BV421-conjugated anti-mouse IFN-γ (Biolegend, Cat. #505830) for 15 min at RT.
Article Title: Enhanced Th1 Cellular Immunity Induced by an RSV-F mRNA Vaccine Rationally Designed Using NLP Algorithms
Article Snippet: .. Lymphocytes were isolated using Mouse 1× Lymphocyte Separation Medium (Dakewe, Cat. #7211011, Dakewe Biotech Co., Ltd., Shenzhen, China), washed twice with PBS, and stimulated with a Respiratory Syncytial Virus Fusion Protein Peptide pool (Vazyme, Cat. #DD9123, 2 μg/mL, Vazyme Biotech Co., Ltd., Nanjing, China) plus CD28 (BD Biosciences, Cat. #553294, BD Biosciences, San Jose, CA, USA) for 18 h. Brefeldin A (BFA) was added and maintained for the final 5 h. First, the cells were stained with a Zombie Aqua Fixable Viability Kit (Biolegend, Cat. #423102, 10 min, RT, BioLegend, Inc., San Diego, CA, USA), followed by APC/Cyanine7 anti-mouse CD3 (Biolegend, Cat. #100330), PerCP anti-mouse CD4 (Biolegend, Cat. #100434), and FITC anti-mouse CD8 (Biolegend, Cat. #100706) for 15–30 min at RT. .. After fixation (Biolegend Fixation Buffer, Cat. #420801) and permeabilization (Biolegend Intracellular Staining Permeabilization Wash Buffer, Cat. #421002), intracellular cytokines were stained with PE-conjugated anti-mouse IL-2 (Biolegend, Cat. #503808), BV605-conjugated anti-mouse IL-4 (Biolegend, Cat. #504126), APC-conjugated anti-mouse TNF-α (Biolegend, Cat. #506308), and BV421-conjugated anti-mouse IFN-γ (Biolegend, Cat. #505830) for 15 min at RT.
|
