Recombinant:Article Title: Highly potent quinoxalinediones inhibit α-hemolysin and ameliorate Staphylococcus aureus lung infections.
Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Recombinant proteins, antibodies and chemicals Alpha-hemolysin IBT Bioservices Cat# 1401-002 Alpha-hemolysin Sigma-Aldrich Cat# H9395 Alpha-hemolysin This study / HZI N/A Mouse anti-Hla mAb IBT Bioservices Cat# 0210-001; RRID: AB_3677702 Rabbit anti-Hla pAb Sigma-Aldrich Cat# S7531; RRID: AB_261550 Histopaque -1077 Sigma Aldrich Cat# 10771 CD14-FITC antibodies Miltenyi Biotec Cat# 130-080-701; RRID: AB_244303 CD14 MicroBeads, human Miltenyi Biotec Cat# 130-050-201 Phorbol 12-myristate 13-acetate Sigma-Aldrich Cat# P1585 VE-Cadherin Antibody Cell Signalling Cat# 2158; RRID: AB_2077970 BiotinPEG4-Azide Jena Bioscience Cat# CLK-1167-5 Streptavidin Sepharose GE Healthcare Cat# 17511301 Critical commercial assays and kits Fluo-4 NW Calcium assay kit Life Technologies Cat# F36206 MultiTox-fluor multiplex cytotoxicity assay Promega Cat# G9200 CellTiter-Glo luminescent viability assay Promega Cat# G7572 LEGENDplex mouse inflammation kit BioLegend Cat# 740446 CytoTox-ONE Homogenous Membrane Integrity assay Promega Cat# G7891 CyQUANT LDH Cytotoxicity assay ThermoFisher Scientific Cat# C20300 FITC Annexin V Apoptosis Detection Kit with PI Biolegend Cat# 640914 NK Cell Isolation Kit, human Miltenyi Biotec Cat# 130-092-657 LIVE/DEAD Fixable Blue Dead Cell Stain Kit Life Technologies Cat# L23105 Colorimetric Sphingomyelinase Assay Kit Sigma-Aldrich Cat# MAK152 Experimental models: cells and blood Buffy coat bags German Red Cross, Springe www.blutspende-nstob.de HSAEpC PromoCell Cat# C-12643 HPMEC PromoCell Cat# C-12281 U937 cells DSMZ ACC5 A549 cells DSMZ ACC107 THP1 cells DSMZ ACC16 Calu-3 cells ATCC HTB-55; RRID: CVCL_0609 Th1 cells CD4+ T cells Miyao et al.49 Rabbit blood Fiebig 36100010 Experimental models: strains and organism CD-1 male mice Charles River N/A C57BL/6 male mice Charles River N/A S. aureus USA300 TCH1516 ATCC BAA-1717 S. aureus USA300 0114 BEI Resources N/A S. aureus USA300 0114 Dhla This study Horsburgh et al.50 and O’Reilly et al.51 Staphylococcus phage 80 Rosenblum and Tyrone52 (Continued on next page) e1 Cell Host & Microbe 33, 1–13.e1–e21, April 9, 2025
Calcium Assay:Article Title: Highly potent quinoxalinediones inhibit α-hemolysin and ameliorate Staphylococcus aureus lung infections.
Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Recombinant proteins, antibodies and chemicals Alpha-hemolysin IBT Bioservices Cat# 1401-002 Alpha-hemolysin Sigma-Aldrich Cat# H9395 Alpha-hemolysin This study / HZI N/A Mouse anti-Hla mAb IBT Bioservices Cat# 0210-001; RRID: AB_3677702 Rabbit anti-Hla pAb Sigma-Aldrich Cat# S7531; RRID: AB_261550 Histopaque -1077 Sigma Aldrich Cat# 10771 CD14-FITC antibodies Miltenyi Biotec Cat# 130-080-701; RRID: AB_244303 CD14 MicroBeads, human Miltenyi Biotec Cat# 130-050-201 Phorbol 12-myristate 13-acetate Sigma-Aldrich Cat# P1585 VE-Cadherin Antibody Cell Signalling Cat# 2158; RRID: AB_2077970 BiotinPEG4-Azide Jena Bioscience Cat# CLK-1167-5 Streptavidin Sepharose GE Healthcare Cat# 17511301 Critical commercial assays and kits Fluo-4 NW Calcium assay kit Life Technologies Cat# F36206 MultiTox-fluor multiplex cytotoxicity assay Promega Cat# G9200 CellTiter-Glo luminescent viability assay Promega Cat# G7572 LEGENDplex mouse inflammation kit BioLegend Cat# 740446 CytoTox-ONE Homogenous Membrane Integrity assay Promega Cat# G7891 CyQUANT LDH Cytotoxicity assay ThermoFisher Scientific Cat# C20300 FITC Annexin V Apoptosis Detection Kit with PI Biolegend Cat# 640914 NK Cell Isolation Kit, human Miltenyi Biotec Cat# 130-092-657 LIVE/DEAD Fixable Blue Dead Cell Stain Kit Life Technologies Cat# L23105 Colorimetric Sphingomyelinase Assay Kit Sigma-Aldrich Cat# MAK152 Experimental models: cells and blood Buffy coat bags German Red Cross, Springe www.blutspende-nstob.de HSAEpC PromoCell Cat# C-12643 HPMEC PromoCell Cat# C-12281 U937 cells DSMZ ACC5 A549 cells DSMZ ACC107 THP1 cells DSMZ ACC16 Calu-3 cells ATCC HTB-55; RRID: CVCL_0609 Th1 cells CD4+ T cells Miyao et al.49 Rabbit blood Fiebig 36100010 Experimental models: strains and organism CD-1 male mice Charles River N/A C57BL/6 male mice Charles River N/A S. aureus USA300 TCH1516 ATCC BAA-1717 S. aureus USA300 0114 BEI Resources N/A S. aureus USA300 0114 Dhla This study Horsburgh et al.50 and O’Reilly et al.51 Staphylococcus phage 80 Rosenblum and Tyrone52 (Continued on next page) e1 Cell Host & Microbe 33, 1–13.e1–e21, April 9, 2025
Multiplex Assay:Article Title: Highly potent quinoxalinediones inhibit α-hemolysin and ameliorate Staphylococcus aureus lung infections.
Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Recombinant proteins, antibodies and chemicals Alpha-hemolysin IBT Bioservices Cat# 1401-002 Alpha-hemolysin Sigma-Aldrich Cat# H9395 Alpha-hemolysin This study / HZI N/A Mouse anti-Hla mAb IBT Bioservices Cat# 0210-001; RRID: AB_3677702 Rabbit anti-Hla pAb Sigma-Aldrich Cat# S7531; RRID: AB_261550 Histopaque -1077 Sigma Aldrich Cat# 10771 CD14-FITC antibodies Miltenyi Biotec Cat# 130-080-701; RRID: AB_244303 CD14 MicroBeads, human Miltenyi Biotec Cat# 130-050-201 Phorbol 12-myristate 13-acetate Sigma-Aldrich Cat# P1585 VE-Cadherin Antibody Cell Signalling Cat# 2158; RRID: AB_2077970 BiotinPEG4-Azide Jena Bioscience Cat# CLK-1167-5 Streptavidin Sepharose GE Healthcare Cat# 17511301 Critical commercial assays and kits Fluo-4 NW Calcium assay kit Life Technologies Cat# F36206 MultiTox-fluor multiplex cytotoxicity assay Promega Cat# G9200 CellTiter-Glo luminescent viability assay Promega Cat# G7572 LEGENDplex mouse inflammation kit BioLegend Cat# 740446 CytoTox-ONE Homogenous Membrane Integrity assay Promega Cat# G7891 CyQUANT LDH Cytotoxicity assay ThermoFisher Scientific Cat# C20300 FITC Annexin V Apoptosis Detection Kit with PI Biolegend Cat# 640914 NK Cell Isolation Kit, human Miltenyi Biotec Cat# 130-092-657 LIVE/DEAD Fixable Blue Dead Cell Stain Kit Life Technologies Cat# L23105 Colorimetric Sphingomyelinase Assay Kit Sigma-Aldrich Cat# MAK152 Experimental models: cells and blood Buffy coat bags German Red Cross, Springe www.blutspende-nstob.de HSAEpC PromoCell Cat# C-12643 HPMEC PromoCell Cat# C-12281 U937 cells DSMZ ACC5 A549 cells DSMZ ACC107 THP1 cells DSMZ ACC16 Calu-3 cells ATCC HTB-55; RRID: CVCL_0609 Th1 cells CD4+ T cells Miyao et al.49 Rabbit blood Fiebig 36100010 Experimental models: strains and organism CD-1 male mice Charles River N/A C57BL/6 male mice Charles River N/A S. aureus USA300 TCH1516 ATCC BAA-1717 S. aureus USA300 0114 BEI Resources N/A S. aureus USA300 0114 Dhla This study Horsburgh et al.50 and O’Reilly et al.51 Staphylococcus phage 80 Rosenblum and Tyrone52 (Continued on next page) e1 Cell Host & Microbe 33, 1–13.e1–e21, April 9, 2025
Cytotoxicity Assay:Article Title: Highly potent quinoxalinediones inhibit α-hemolysin and ameliorate Staphylococcus aureus lung infections.
Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Recombinant proteins, antibodies and chemicals Alpha-hemolysin IBT Bioservices Cat# 1401-002 Alpha-hemolysin Sigma-Aldrich Cat# H9395 Alpha-hemolysin This study / HZI N/A Mouse anti-Hla mAb IBT Bioservices Cat# 0210-001; RRID: AB_3677702 Rabbit anti-Hla pAb Sigma-Aldrich Cat# S7531; RRID: AB_261550 Histopaque -1077 Sigma Aldrich Cat# 10771 CD14-FITC antibodies Miltenyi Biotec Cat# 130-080-701; RRID: AB_244303 CD14 MicroBeads, human Miltenyi Biotec Cat# 130-050-201 Phorbol 12-myristate 13-acetate Sigma-Aldrich Cat# P1585 VE-Cadherin Antibody Cell Signalling Cat# 2158; RRID: AB_2077970 BiotinPEG4-Azide Jena Bioscience Cat# CLK-1167-5 Streptavidin Sepharose GE Healthcare Cat# 17511301 Critical commercial assays and kits Fluo-4 NW Calcium assay kit Life Technologies Cat# F36206 MultiTox-fluor multiplex cytotoxicity assay Promega Cat# G9200 CellTiter-Glo luminescent viability assay Promega Cat# G7572 LEGENDplex mouse inflammation kit BioLegend Cat# 740446 CytoTox-ONE Homogenous Membrane Integrity assay Promega Cat# G7891 CyQUANT LDH Cytotoxicity assay ThermoFisher Scientific Cat# C20300 FITC Annexin V Apoptosis Detection Kit with PI Biolegend Cat# 640914 NK Cell Isolation Kit, human Miltenyi Biotec Cat# 130-092-657 LIVE/DEAD Fixable Blue Dead Cell Stain Kit Life Technologies Cat# L23105 Colorimetric Sphingomyelinase Assay Kit Sigma-Aldrich Cat# MAK152 Experimental models: cells and blood Buffy coat bags German Red Cross, Springe www.blutspende-nstob.de HSAEpC PromoCell Cat# C-12643 HPMEC PromoCell Cat# C-12281 U937 cells DSMZ ACC5 A549 cells DSMZ ACC107 THP1 cells DSMZ ACC16 Calu-3 cells ATCC HTB-55; RRID: CVCL_0609 Th1 cells CD4+ T cells Miyao et al.49 Rabbit blood Fiebig 36100010 Experimental models: strains and organism CD-1 male mice Charles River N/A C57BL/6 male mice Charles River N/A S. aureus USA300 TCH1516 ATCC BAA-1717 S. aureus USA300 0114 BEI Resources N/A S. aureus USA300 0114 Dhla This study Horsburgh et al.50 and O’Reilly et al.51 Staphylococcus phage 80 Rosenblum and Tyrone52 (Continued on next page) e1 Cell Host & Microbe 33, 1–13.e1–e21, April 9, 2025
Viability Assay:Article Title: Highly potent quinoxalinediones inhibit α-hemolysin and ameliorate Staphylococcus aureus lung infections.
Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Recombinant proteins, antibodies and chemicals Alpha-hemolysin IBT Bioservices Cat# 1401-002 Alpha-hemolysin Sigma-Aldrich Cat# H9395 Alpha-hemolysin This study / HZI N/A Mouse anti-Hla mAb IBT Bioservices Cat# 0210-001; RRID: AB_3677702 Rabbit anti-Hla pAb Sigma-Aldrich Cat# S7531; RRID: AB_261550 Histopaque -1077 Sigma Aldrich Cat# 10771 CD14-FITC antibodies Miltenyi Biotec Cat# 130-080-701; RRID: AB_244303 CD14 MicroBeads, human Miltenyi Biotec Cat# 130-050-201 Phorbol 12-myristate 13-acetate Sigma-Aldrich Cat# P1585 VE-Cadherin Antibody Cell Signalling Cat# 2158; RRID: AB_2077970 BiotinPEG4-Azide Jena Bioscience Cat# CLK-1167-5 Streptavidin Sepharose GE Healthcare Cat# 17511301 Critical commercial assays and kits Fluo-4 NW Calcium assay kit Life Technologies Cat# F36206 MultiTox-fluor multiplex cytotoxicity assay Promega Cat# G9200 CellTiter-Glo luminescent viability assay Promega Cat# G7572 LEGENDplex mouse inflammation kit BioLegend Cat# 740446 CytoTox-ONE Homogenous Membrane Integrity assay Promega Cat# G7891 CyQUANT LDH Cytotoxicity assay ThermoFisher Scientific Cat# C20300 FITC Annexin V Apoptosis Detection Kit with PI Biolegend Cat# 640914 NK Cell Isolation Kit, human Miltenyi Biotec Cat# 130-092-657 LIVE/DEAD Fixable Blue Dead Cell Stain Kit Life Technologies Cat# L23105 Colorimetric Sphingomyelinase Assay Kit Sigma-Aldrich Cat# MAK152 Experimental models: cells and blood Buffy coat bags German Red Cross, Springe www.blutspende-nstob.de HSAEpC PromoCell Cat# C-12643 HPMEC PromoCell Cat# C-12281 U937 cells DSMZ ACC5 A549 cells DSMZ ACC107 THP1 cells DSMZ ACC16 Calu-3 cells ATCC HTB-55; RRID: CVCL_0609 Th1 cells CD4+ T cells Miyao et al.49 Rabbit blood Fiebig 36100010 Experimental models: strains and organism CD-1 male mice Charles River N/A C57BL/6 male mice Charles River N/A S. aureus USA300 TCH1516 ATCC BAA-1717 S. aureus USA300 0114 BEI Resources N/A S. aureus USA300 0114 Dhla This study Horsburgh et al.50 and O’Reilly et al.51 Staphylococcus phage 80 Rosenblum and Tyrone52 (Continued on next page) e1 Cell Host & Microbe 33, 1–13.e1–e21, April 9, 2025
Membrane:Article Title: Highly potent quinoxalinediones inhibit α-hemolysin and ameliorate Staphylococcus aureus lung infections.
Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Recombinant proteins, antibodies and chemicals Alpha-hemolysin IBT Bioservices Cat# 1401-002 Alpha-hemolysin Sigma-Aldrich Cat# H9395 Alpha-hemolysin This study / HZI N/A Mouse anti-Hla mAb IBT Bioservices Cat# 0210-001; RRID: AB_3677702 Rabbit anti-Hla pAb Sigma-Aldrich Cat# S7531; RRID: AB_261550 Histopaque -1077 Sigma Aldrich Cat# 10771 CD14-FITC antibodies Miltenyi Biotec Cat# 130-080-701; RRID: AB_244303 CD14 MicroBeads, human Miltenyi Biotec Cat# 130-050-201 Phorbol 12-myristate 13-acetate Sigma-Aldrich Cat# P1585 VE-Cadherin Antibody Cell Signalling Cat# 2158; RRID: AB_2077970 BiotinPEG4-Azide Jena Bioscience Cat# CLK-1167-5 Streptavidin Sepharose GE Healthcare Cat# 17511301 Critical commercial assays and kits Fluo-4 NW Calcium assay kit Life Technologies Cat# F36206 MultiTox-fluor multiplex cytotoxicity assay Promega Cat# G9200 CellTiter-Glo luminescent viability assay Promega Cat# G7572 LEGENDplex mouse inflammation kit BioLegend Cat# 740446 CytoTox-ONE Homogenous Membrane Integrity assay Promega Cat# G7891 CyQUANT LDH Cytotoxicity assay ThermoFisher Scientific Cat# C20300 FITC Annexin V Apoptosis Detection Kit with PI Biolegend Cat# 640914 NK Cell Isolation Kit, human Miltenyi Biotec Cat# 130-092-657 LIVE/DEAD Fixable Blue Dead Cell Stain Kit Life Technologies Cat# L23105 Colorimetric Sphingomyelinase Assay Kit Sigma-Aldrich Cat# MAK152 Experimental models: cells and blood Buffy coat bags German Red Cross, Springe www.blutspende-nstob.de HSAEpC PromoCell Cat# C-12643 HPMEC PromoCell Cat# C-12281 U937 cells DSMZ ACC5 A549 cells DSMZ ACC107 THP1 cells DSMZ ACC16 Calu-3 cells ATCC HTB-55; RRID: CVCL_0609 Th1 cells CD4+ T cells Miyao et al.49 Rabbit blood Fiebig 36100010 Experimental models: strains and organism CD-1 male mice Charles River N/A C57BL/6 male mice Charles River N/A S. aureus USA300 TCH1516 ATCC BAA-1717 S. aureus USA300 0114 BEI Resources N/A S. aureus USA300 0114 Dhla This study Horsburgh et al.50 and O’Reilly et al.51 Staphylococcus phage 80 Rosenblum and Tyrone52 (Continued on next page) e1 Cell Host & Microbe 33, 1–13.e1–e21, April 9, 2025
Integrity Assay:Article Title: Highly potent quinoxalinediones inhibit α-hemolysin and ameliorate Staphylococcus aureus lung infections.
Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Recombinant proteins, antibodies and chemicals Alpha-hemolysin IBT Bioservices Cat# 1401-002 Alpha-hemolysin Sigma-Aldrich Cat# H9395 Alpha-hemolysin This study / HZI N/A Mouse anti-Hla mAb IBT Bioservices Cat# 0210-001; RRID: AB_3677702 Rabbit anti-Hla pAb Sigma-Aldrich Cat# S7531; RRID: AB_261550 Histopaque -1077 Sigma Aldrich Cat# 10771 CD14-FITC antibodies Miltenyi Biotec Cat# 130-080-701; RRID: AB_244303 CD14 MicroBeads, human Miltenyi Biotec Cat# 130-050-201 Phorbol 12-myristate 13-acetate Sigma-Aldrich Cat# P1585 VE-Cadherin Antibody Cell Signalling Cat# 2158; RRID: AB_2077970 BiotinPEG4-Azide Jena Bioscience Cat# CLK-1167-5 Streptavidin Sepharose GE Healthcare Cat# 17511301 Critical commercial assays and kits Fluo-4 NW Calcium assay kit Life Technologies Cat# F36206 MultiTox-fluor multiplex cytotoxicity assay Promega Cat# G9200 CellTiter-Glo luminescent viability assay Promega Cat# G7572 LEGENDplex mouse inflammation kit BioLegend Cat# 740446 CytoTox-ONE Homogenous Membrane Integrity assay Promega Cat# G7891 CyQUANT LDH Cytotoxicity assay ThermoFisher Scientific Cat# C20300 FITC Annexin V Apoptosis Detection Kit with PI Biolegend Cat# 640914 NK Cell Isolation Kit, human Miltenyi Biotec Cat# 130-092-657 LIVE/DEAD Fixable Blue Dead Cell Stain Kit Life Technologies Cat# L23105 Colorimetric Sphingomyelinase Assay Kit Sigma-Aldrich Cat# MAK152 Experimental models: cells and blood Buffy coat bags German Red Cross, Springe www.blutspende-nstob.de HSAEpC PromoCell Cat# C-12643 HPMEC PromoCell Cat# C-12281 U937 cells DSMZ ACC5 A549 cells DSMZ ACC107 THP1 cells DSMZ ACC16 Calu-3 cells ATCC HTB-55; RRID: CVCL_0609 Th1 cells CD4+ T cells Miyao et al.49 Rabbit blood Fiebig 36100010 Experimental models: strains and organism CD-1 male mice Charles River N/A C57BL/6 male mice Charles River N/A S. aureus USA300 TCH1516 ATCC BAA-1717 S. aureus USA300 0114 BEI Resources N/A S. aureus USA300 0114 Dhla This study Horsburgh et al.50 and O’Reilly et al.51 Staphylococcus phage 80 Rosenblum and Tyrone52 (Continued on next page) e1 Cell Host & Microbe 33, 1–13.e1–e21, April 9, 2025
CyQUANT Assay:Article Title: Highly potent quinoxalinediones inhibit α-hemolysin and ameliorate Staphylococcus aureus lung infections.
Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Recombinant proteins, antibodies and chemicals Alpha-hemolysin IBT Bioservices Cat# 1401-002 Alpha-hemolysin Sigma-Aldrich Cat# H9395 Alpha-hemolysin This study / HZI N/A Mouse anti-Hla mAb IBT Bioservices Cat# 0210-001; RRID: AB_3677702 Rabbit anti-Hla pAb Sigma-Aldrich Cat# S7531; RRID: AB_261550 Histopaque -1077 Sigma Aldrich Cat# 10771 CD14-FITC antibodies Miltenyi Biotec Cat# 130-080-701; RRID: AB_244303 CD14 MicroBeads, human Miltenyi Biotec Cat# 130-050-201 Phorbol 12-myristate 13-acetate Sigma-Aldrich Cat# P1585 VE-Cadherin Antibody Cell Signalling Cat# 2158; RRID: AB_2077970 BiotinPEG4-Azide Jena Bioscience Cat# CLK-1167-5 Streptavidin Sepharose GE Healthcare Cat# 17511301 Critical commercial assays and kits Fluo-4 NW Calcium assay kit Life Technologies Cat# F36206 MultiTox-fluor multiplex cytotoxicity assay Promega Cat# G9200 CellTiter-Glo luminescent viability assay Promega Cat# G7572 LEGENDplex mouse inflammation kit BioLegend Cat# 740446 CytoTox-ONE Homogenous Membrane Integrity assay Promega Cat# G7891 CyQUANT LDH Cytotoxicity assay ThermoFisher Scientific Cat# C20300 FITC Annexin V Apoptosis Detection Kit with PI Biolegend Cat# 640914 NK Cell Isolation Kit, human Miltenyi Biotec Cat# 130-092-657 LIVE/DEAD Fixable Blue Dead Cell Stain Kit Life Technologies Cat# L23105 Colorimetric Sphingomyelinase Assay Kit Sigma-Aldrich Cat# MAK152 Experimental models: cells and blood Buffy coat bags German Red Cross, Springe www.blutspende-nstob.de HSAEpC PromoCell Cat# C-12643 HPMEC PromoCell Cat# C-12281 U937 cells DSMZ ACC5 A549 cells DSMZ ACC107 THP1 cells DSMZ ACC16 Calu-3 cells ATCC HTB-55; RRID: CVCL_0609 Th1 cells CD4+ T cells Miyao et al.49 Rabbit blood Fiebig 36100010 Experimental models: strains and organism CD-1 male mice Charles River N/A C57BL/6 male mice Charles River N/A S. aureus USA300 TCH1516 ATCC BAA-1717 S. aureus USA300 0114 BEI Resources N/A S. aureus USA300 0114 Dhla This study Horsburgh et al.50 and O’Reilly et al.51 Staphylococcus phage 80 Rosenblum and Tyrone52 (Continued on next page) e1 Cell Host & Microbe 33, 1–13.e1–e21, April 9, 2025
LDH Cytotoxicity Assay:Article Title: Highly potent quinoxalinediones inhibit α-hemolysin and ameliorate Staphylococcus aureus lung infections.
Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Recombinant proteins, antibodies and chemicals Alpha-hemolysin IBT Bioservices Cat# 1401-002 Alpha-hemolysin Sigma-Aldrich Cat# H9395 Alpha-hemolysin This study / HZI N/A Mouse anti-Hla mAb IBT Bioservices Cat# 0210-001; RRID: AB_3677702 Rabbit anti-Hla pAb Sigma-Aldrich Cat# S7531; RRID: AB_261550 Histopaque -1077 Sigma Aldrich Cat# 10771 CD14-FITC antibodies Miltenyi Biotec Cat# 130-080-701; RRID: AB_244303 CD14 MicroBeads, human Miltenyi Biotec Cat# 130-050-201 Phorbol 12-myristate 13-acetate Sigma-Aldrich Cat# P1585 VE-Cadherin Antibody Cell Signalling Cat# 2158; RRID: AB_2077970 BiotinPEG4-Azide Jena Bioscience Cat# CLK-1167-5 Streptavidin Sepharose GE Healthcare Cat# 17511301 Critical commercial assays and kits Fluo-4 NW Calcium assay kit Life Technologies Cat# F36206 MultiTox-fluor multiplex cytotoxicity assay Promega Cat# G9200 CellTiter-Glo luminescent viability assay Promega Cat# G7572 LEGENDplex mouse inflammation kit BioLegend Cat# 740446 CytoTox-ONE Homogenous Membrane Integrity assay Promega Cat# G7891 CyQUANT LDH Cytotoxicity assay ThermoFisher Scientific Cat# C20300 FITC Annexin V Apoptosis Detection Kit with PI Biolegend Cat# 640914 NK Cell Isolation Kit, human Miltenyi Biotec Cat# 130-092-657 LIVE/DEAD Fixable Blue Dead Cell Stain Kit Life Technologies Cat# L23105 Colorimetric Sphingomyelinase Assay Kit Sigma-Aldrich Cat# MAK152 Experimental models: cells and blood Buffy coat bags German Red Cross, Springe www.blutspende-nstob.de HSAEpC PromoCell Cat# C-12643 HPMEC PromoCell Cat# C-12281 U937 cells DSMZ ACC5 A549 cells DSMZ ACC107 THP1 cells DSMZ ACC16 Calu-3 cells ATCC HTB-55; RRID: CVCL_0609 Th1 cells CD4+ T cells Miyao et al.49 Rabbit blood Fiebig 36100010 Experimental models: strains and organism CD-1 male mice Charles River N/A C57BL/6 male mice Charles River N/A S. aureus USA300 TCH1516 ATCC BAA-1717 S. aureus USA300 0114 BEI Resources N/A S. aureus USA300 0114 Dhla This study Horsburgh et al.50 and O’Reilly et al.51 Staphylococcus phage 80 Rosenblum and Tyrone52 (Continued on next page) e1 Cell Host & Microbe 33, 1–13.e1–e21, April 9, 2025
Cell Isolation:Article Title: Highly potent quinoxalinediones inhibit α-hemolysin and ameliorate Staphylococcus aureus lung infections.
Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Recombinant proteins, antibodies and chemicals Alpha-hemolysin IBT Bioservices Cat# 1401-002 Alpha-hemolysin Sigma-Aldrich Cat# H9395 Alpha-hemolysin This study / HZI N/A Mouse anti-Hla mAb IBT Bioservices Cat# 0210-001; RRID: AB_3677702 Rabbit anti-Hla pAb Sigma-Aldrich Cat# S7531; RRID: AB_261550 Histopaque -1077 Sigma Aldrich Cat# 10771 CD14-FITC antibodies Miltenyi Biotec Cat# 130-080-701; RRID: AB_244303 CD14 MicroBeads, human Miltenyi Biotec Cat# 130-050-201 Phorbol 12-myristate 13-acetate Sigma-Aldrich Cat# P1585 VE-Cadherin Antibody Cell Signalling Cat# 2158; RRID: AB_2077970 BiotinPEG4-Azide Jena Bioscience Cat# CLK-1167-5 Streptavidin Sepharose GE Healthcare Cat# 17511301 Critical commercial assays and kits Fluo-4 NW Calcium assay kit Life Technologies Cat# F36206 MultiTox-fluor multiplex cytotoxicity assay Promega Cat# G9200 CellTiter-Glo luminescent viability assay Promega Cat# G7572 LEGENDplex mouse inflammation kit BioLegend Cat# 740446 CytoTox-ONE Homogenous Membrane Integrity assay Promega Cat# G7891 CyQUANT LDH Cytotoxicity assay ThermoFisher Scientific Cat# C20300 FITC Annexin V Apoptosis Detection Kit with PI Biolegend Cat# 640914 NK Cell Isolation Kit, human Miltenyi Biotec Cat# 130-092-657 LIVE/DEAD Fixable Blue Dead Cell Stain Kit Life Technologies Cat# L23105 Colorimetric Sphingomyelinase Assay Kit Sigma-Aldrich Cat# MAK152 Experimental models: cells and blood Buffy coat bags German Red Cross, Springe www.blutspende-nstob.de HSAEpC PromoCell Cat# C-12643 HPMEC PromoCell Cat# C-12281 U937 cells DSMZ ACC5 A549 cells DSMZ ACC107 THP1 cells DSMZ ACC16 Calu-3 cells ATCC HTB-55; RRID: CVCL_0609 Th1 cells CD4+ T cells Miyao et al.49 Rabbit blood Fiebig 36100010 Experimental models: strains and organism CD-1 male mice Charles River N/A C57BL/6 male mice Charles River N/A S. aureus USA300 TCH1516 ATCC BAA-1717 S. aureus USA300 0114 BEI Resources N/A S. aureus USA300 0114 Dhla This study Horsburgh et al.50 and O’Reilly et al.51 Staphylococcus phage 80 Rosenblum and Tyrone52 (Continued on next page) e1 Cell Host & Microbe 33, 1–13.e1–e21, April 9, 2025
Staining:Article Title: Highly potent quinoxalinediones inhibit α-hemolysin and ameliorate Staphylococcus aureus lung infections.
Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Recombinant proteins, antibodies and chemicals Alpha-hemolysin IBT Bioservices Cat# 1401-002 Alpha-hemolysin Sigma-Aldrich Cat# H9395 Alpha-hemolysin This study / HZI N/A Mouse anti-Hla mAb IBT Bioservices Cat# 0210-001; RRID: AB_3677702 Rabbit anti-Hla pAb Sigma-Aldrich Cat# S7531; RRID: AB_261550 Histopaque -1077 Sigma Aldrich Cat# 10771 CD14-FITC antibodies Miltenyi Biotec Cat# 130-080-701; RRID: AB_244303 CD14 MicroBeads, human Miltenyi Biotec Cat# 130-050-201 Phorbol 12-myristate 13-acetate Sigma-Aldrich Cat# P1585 VE-Cadherin Antibody Cell Signalling Cat# 2158; RRID: AB_2077970 BiotinPEG4-Azide Jena Bioscience Cat# CLK-1167-5 Streptavidin Sepharose GE Healthcare Cat# 17511301 Critical commercial assays and kits Fluo-4 NW Calcium assay kit Life Technologies Cat# F36206 MultiTox-fluor multiplex cytotoxicity assay Promega Cat# G9200 CellTiter-Glo luminescent viability assay Promega Cat# G7572 LEGENDplex mouse inflammation kit BioLegend Cat# 740446 CytoTox-ONE Homogenous Membrane Integrity assay Promega Cat# G7891 CyQUANT LDH Cytotoxicity assay ThermoFisher Scientific Cat# C20300 FITC Annexin V Apoptosis Detection Kit with PI Biolegend Cat# 640914 NK Cell Isolation Kit, human Miltenyi Biotec Cat# 130-092-657 LIVE/DEAD Fixable Blue Dead Cell Stain Kit Life Technologies Cat# L23105 Colorimetric Sphingomyelinase Assay Kit Sigma-Aldrich Cat# MAK152 Experimental models: cells and blood Buffy coat bags German Red Cross, Springe www.blutspende-nstob.de HSAEpC PromoCell Cat# C-12643 HPMEC PromoCell Cat# C-12281 U937 cells DSMZ ACC5 A549 cells DSMZ ACC107 THP1 cells DSMZ ACC16 Calu-3 cells ATCC HTB-55; RRID: CVCL_0609 Th1 cells CD4+ T cells Miyao et al.49 Rabbit blood Fiebig 36100010 Experimental models: strains and organism CD-1 male mice Charles River N/A C57BL/6 male mice Charles River N/A S. aureus USA300 TCH1516 ATCC BAA-1717 S. aureus USA300 0114 BEI Resources N/A S. aureus USA300 0114 Dhla This study Horsburgh et al.50 and O’Reilly et al.51 Staphylococcus phage 80 Rosenblum and Tyrone52 (Continued on next page) e1 Cell Host & Microbe 33, 1–13.e1–e21, April 9, 2025
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