Isolation:Article Title: NET formation-mediated in situ protein delivery to the inflamed central nervous system.
Article Snippet: .. For astrocyte isolation, spinal cord single-cell suspensions were stained with the following antibodies for sorting: CD45R/B220 PE (1:200, cat# 103207, lot# B411979, clone RA3-6B2, BioLegend), TER-119 PE (1:200, cat# 116207, lot# B392792, clone TER119, BioLegend), O4 PE (1:100, cat# 130-117-507, lot# 5240701123, clone O4,Miltenyi Biotec), CD105 PE (1:200, cat# 120407, lot# B364575, clone MJ7/18, BioLegend), CD140a PE (1:200, cat# 135905, lot# B401844, clone APA5, BioLegend), Ly6G PE (1:200, cat# 164503, lot# B417691, clone S19018G, BioLegend), Ly6C APC/Cy7 (1:200, cat# 128025, lot# B417977, clone HK1.4, BioLegend), CD45 Alexa Fluor 647 (1:100, cat# 103124, lot# B364327, clone 30-F11, BioLegend), CD11c FITC (1:100, cat# 117306, lot# B353146, clone N418, BioLegend), andCD11b BV605 (1:100, cat# 101257, lot# B363316, clone M1/70, BioLegend). .. Astrocytes were gated asTer119-O4-CD45R-CD140a-CD105-Ly6G-CD45-CD11b-CD11c-Ly6Cand sorted using a FACSAria III (BD Biosciences).
Article Title: NET formation-mediated in situ protein delivery to the inflamed central nervous system
Article Snippet: .. For astrocyte isolation, spinal cord single-cell suspensions were stained with the following antibodies for sorting: CD45R/B220 PE (1:200, cat# 103207, lot# B411979, clone RA3-6B2, BioLegend), TER-119 PE (1:200, cat# 116207, lot# B392792, clone TER-119, BioLegend), O4 PE (1:100, cat# 130-117-507, lot# 5240701123, clone O4, Miltenyi Biotec), CD105 PE (1:200, cat# 120407, lot# B364575, clone MJ7/18, BioLegend), CD140a PE (1:200, cat# 135905, lot# B401844, clone APA5, BioLegend), Ly6G PE (1:200, cat# 164503, lot# B417691, clone S19018G, BioLegend), Ly6C APC/Cy7 (1:200, cat# 128025, lot# B417977, clone HK1.4, BioLegend), CD45 Alexa Fluor 647 (1:100, cat# 103124, lot# B364327, clone 30-F11, BioLegend), CD11c FITC (1:100, cat# 117306, lot# B353146, clone N418, BioLegend), and CD11b BV605 (1:100, cat# 101257, lot# B363316, clone M1/70, BioLegend). .. Astrocytes were gated as Ter119 - O4 - CD45R - CD140a - CD105 - Ly6G - CD45 - CD11b - CD11c - Ly6C - and sorted using a FACSAria III (BD Biosciences).
Single Cell:Article Title: NET formation-mediated in situ protein delivery to the inflamed central nervous system.
Article Snippet: .. For astrocyte isolation, spinal cord single-cell suspensions were stained with the following antibodies for sorting: CD45R/B220 PE (1:200, cat# 103207, lot# B411979, clone RA3-6B2, BioLegend), TER-119 PE (1:200, cat# 116207, lot# B392792, clone TER119, BioLegend), O4 PE (1:100, cat# 130-117-507, lot# 5240701123, clone O4,Miltenyi Biotec), CD105 PE (1:200, cat# 120407, lot# B364575, clone MJ7/18, BioLegend), CD140a PE (1:200, cat# 135905, lot# B401844, clone APA5, BioLegend), Ly6G PE (1:200, cat# 164503, lot# B417691, clone S19018G, BioLegend), Ly6C APC/Cy7 (1:200, cat# 128025, lot# B417977, clone HK1.4, BioLegend), CD45 Alexa Fluor 647 (1:100, cat# 103124, lot# B364327, clone 30-F11, BioLegend), CD11c FITC (1:100, cat# 117306, lot# B353146, clone N418, BioLegend), andCD11b BV605 (1:100, cat# 101257, lot# B363316, clone M1/70, BioLegend). .. Astrocytes were gated asTer119-O4-CD45R-CD140a-CD105-Ly6G-CD45-CD11b-CD11c-Ly6Cand sorted using a FACSAria III (BD Biosciences).
Article Title: NET formation-mediated in situ protein delivery to the inflamed central nervous system
Article Snippet: .. For astrocyte isolation, spinal cord single-cell suspensions were stained with the following antibodies for sorting: CD45R/B220 PE (1:200, cat# 103207, lot# B411979, clone RA3-6B2, BioLegend), TER-119 PE (1:200, cat# 116207, lot# B392792, clone TER-119, BioLegend), O4 PE (1:100, cat# 130-117-507, lot# 5240701123, clone O4, Miltenyi Biotec), CD105 PE (1:200, cat# 120407, lot# B364575, clone MJ7/18, BioLegend), CD140a PE (1:200, cat# 135905, lot# B401844, clone APA5, BioLegend), Ly6G PE (1:200, cat# 164503, lot# B417691, clone S19018G, BioLegend), Ly6C APC/Cy7 (1:200, cat# 128025, lot# B417977, clone HK1.4, BioLegend), CD45 Alexa Fluor 647 (1:100, cat# 103124, lot# B364327, clone 30-F11, BioLegend), CD11c FITC (1:100, cat# 117306, lot# B353146, clone N418, BioLegend), and CD11b BV605 (1:100, cat# 101257, lot# B363316, clone M1/70, BioLegend). .. Astrocytes were gated as Ter119 - O4 - CD45R - CD140a - CD105 - Ly6G - CD45 - CD11b - CD11c - Ly6C - and sorted using a FACSAria III (BD Biosciences).
Staining:Article Title: NET formation-mediated in situ protein delivery to the inflamed central nervous system.
Article Snippet: .. For astrocyte isolation, spinal cord single-cell suspensions were stained with the following antibodies for sorting: CD45R/B220 PE (1:200, cat# 103207, lot# B411979, clone RA3-6B2, BioLegend), TER-119 PE (1:200, cat# 116207, lot# B392792, clone TER119, BioLegend), O4 PE (1:100, cat# 130-117-507, lot# 5240701123, clone O4,Miltenyi Biotec), CD105 PE (1:200, cat# 120407, lot# B364575, clone MJ7/18, BioLegend), CD140a PE (1:200, cat# 135905, lot# B401844, clone APA5, BioLegend), Ly6G PE (1:200, cat# 164503, lot# B417691, clone S19018G, BioLegend), Ly6C APC/Cy7 (1:200, cat# 128025, lot# B417977, clone HK1.4, BioLegend), CD45 Alexa Fluor 647 (1:100, cat# 103124, lot# B364327, clone 30-F11, BioLegend), CD11c FITC (1:100, cat# 117306, lot# B353146, clone N418, BioLegend), andCD11b BV605 (1:100, cat# 101257, lot# B363316, clone M1/70, BioLegend). .. Astrocytes were gated asTer119-O4-CD45R-CD140a-CD105-Ly6G-CD45-CD11b-CD11c-Ly6Cand sorted using a FACSAria III (BD Biosciences).
Article Title: NET formation-mediated in situ protein delivery to the inflamed central nervous system
Article Snippet: .. For astrocyte isolation, spinal cord single-cell suspensions were stained with the following antibodies for sorting: CD45R/B220 PE (1:200, cat# 103207, lot# B411979, clone RA3-6B2, BioLegend), TER-119 PE (1:200, cat# 116207, lot# B392792, clone TER-119, BioLegend), O4 PE (1:100, cat# 130-117-507, lot# 5240701123, clone O4, Miltenyi Biotec), CD105 PE (1:200, cat# 120407, lot# B364575, clone MJ7/18, BioLegend), CD140a PE (1:200, cat# 135905, lot# B401844, clone APA5, BioLegend), Ly6G PE (1:200, cat# 164503, lot# B417691, clone S19018G, BioLegend), Ly6C APC/Cy7 (1:200, cat# 128025, lot# B417977, clone HK1.4, BioLegend), CD45 Alexa Fluor 647 (1:100, cat# 103124, lot# B364327, clone 30-F11, BioLegend), CD11c FITC (1:100, cat# 117306, lot# B353146, clone N418, BioLegend), and CD11b BV605 (1:100, cat# 101257, lot# B363316, clone M1/70, BioLegend). .. Astrocytes were gated as Ter119 - O4 - CD45R - CD140a - CD105 - Ly6G - CD45 - CD11b - CD11c - Ly6C - and sorted using a FACSAria III (BD Biosciences).
Incubation:Article Title: Systemic inhibition of soluble TNF significantly changes glial cell populations leading to improved myelin integrity and better functional outcome after experimental stroke
Article Snippet: .. Glial cell samples were incubated with CD45-PerCP-Cy 5.5 (clone: 30-F11; BD Biosciences, #561869, diluted 1:100), astrocyte cell surface antigen-2 (ACSA-2)-FITC (clone: REA969; Miltenyi Biotec, #130–116–243, diluted 1:50), CD140a-APC (PDGFRα; clone: REA637; Miltenyi Biotec, #130–122–942, diluted 1:10), and O4-PE (clone: REA576; Miltenyi Biotec, #130–117–823, diluted 1:50), while immune cell samples were incubated with CD45-PerCP-Cy 5.5 (clone: 30-F11), CD11b-BB515 (clone: M1/70; BD Biosciences, #564455, diluted 1:200), CD4-APC (clone: RM4–5; BD Biosciences, #561091, diluted 1:200), CD8a-PE (clone: 53–6.7; BD Biosciences #553032, diluted 1:200), Lymphocyte antigen 6 G (Ly-6G)-BV786 (clone: 1A8; BD Biosciences, #740953, diluted 1:200), and Major histocompatibility complex II (MHCII)-APC-Cy7 (clone: M5/114.15.2; Biolegend, #107627, diluted 1:200). .. All samples were fixed in 1 % PFA, and Trucount counting beads (5000 beads/sample; BD Trucount Absolute Counting Tubes, BD Biosciences, #663028) were added for use in estimation of total cell numbers.
Article Title: Systemic inhibition of soluble TNF significantly changes glial cell populations leading to improved myelin integrity and better functional outcome after experimental stroke.
Article Snippet: .. Glial cell samples were incubated with CD45-PerCP-Cy5.5 (clone: 30-F11; BD Biosciences, #561869, diluted 1:100), astrocyte cell surface antigen-2 (ACSA-2)-FITC (clone: REA969; Miltenyi Biotec, #130–116–243, diluted 1:50), CD140a-APC (PDGFRα; clone: REA637; Miltenyi Biotec, #130–122–942, diluted 1:10), and O4-PE (clone: REA576; Miltenyi Biotec, #130–117–823, diluted 1:50), while immune cell samples were incubated with CD45-PerCP-Cy5.5 (clone: 30-F11), CD11b-BB515 (clone: M1/70; BD Biosciences, #564455, diluted 1:200), CD4-APC (clone: RM4–5; BD Biosciences, #561091, diluted 1:200), CD8a-PE (clone: 53–6.7; BD Biosciences #553032, diluted 1:200), Lymphocyte antigen 6 G (Ly-6G)-BV786 (clone: 1A8; BD Biosciences, #740953, diluted 1:200), and Major histocompatibility complex II (MHCII)-APC-Cy7 (clone: M5/114.15.2; Biolegend, #107627, diluted 1:200). .. All samples were fixed in 1 % PFA, and Trucount counting beads (5000 beads/sample; BD Trucount Absolute Counting Tubes, BD Biosciences, #663028) were added for use in estimation of total cell numbers.
Immune Cell:Article Title: Systemic inhibition of soluble TNF significantly changes glial cell populations leading to improved myelin integrity and better functional outcome after experimental stroke
Article Snippet: .. Glial cell samples were incubated with CD45-PerCP-Cy 5.5 (clone: 30-F11; BD Biosciences, #561869, diluted 1:100), astrocyte cell surface antigen-2 (ACSA-2)-FITC (clone: REA969; Miltenyi Biotec, #130–116–243, diluted 1:50), CD140a-APC (PDGFRα; clone: REA637; Miltenyi Biotec, #130–122–942, diluted 1:10), and O4-PE (clone: REA576; Miltenyi Biotec, #130–117–823, diluted 1:50), while immune cell samples were incubated with CD45-PerCP-Cy 5.5 (clone: 30-F11), CD11b-BB515 (clone: M1/70; BD Biosciences, #564455, diluted 1:200), CD4-APC (clone: RM4–5; BD Biosciences, #561091, diluted 1:200), CD8a-PE (clone: 53–6.7; BD Biosciences #553032, diluted 1:200), Lymphocyte antigen 6 G (Ly-6G)-BV786 (clone: 1A8; BD Biosciences, #740953, diluted 1:200), and Major histocompatibility complex II (MHCII)-APC-Cy7 (clone: M5/114.15.2; Biolegend, #107627, diluted 1:200). .. All samples were fixed in 1 % PFA, and Trucount counting beads (5000 beads/sample; BD Trucount Absolute Counting Tubes, BD Biosciences, #663028) were added for use in estimation of total cell numbers.
Article Title: Systemic inhibition of soluble TNF significantly changes glial cell populations leading to improved myelin integrity and better functional outcome after experimental stroke.
Article Snippet: .. Glial cell samples were incubated with CD45-PerCP-Cy5.5 (clone: 30-F11; BD Biosciences, #561869, diluted 1:100), astrocyte cell surface antigen-2 (ACSA-2)-FITC (clone: REA969; Miltenyi Biotec, #130–116–243, diluted 1:50), CD140a-APC (PDGFRα; clone: REA637; Miltenyi Biotec, #130–122–942, diluted 1:10), and O4-PE (clone: REA576; Miltenyi Biotec, #130–117–823, diluted 1:50), while immune cell samples were incubated with CD45-PerCP-Cy5.5 (clone: 30-F11), CD11b-BB515 (clone: M1/70; BD Biosciences, #564455, diluted 1:200), CD4-APC (clone: RM4–5; BD Biosciences, #561091, diluted 1:200), CD8a-PE (clone: 53–6.7; BD Biosciences #553032, diluted 1:200), Lymphocyte antigen 6 G (Ly-6G)-BV786 (clone: 1A8; BD Biosciences, #740953, diluted 1:200), and Major histocompatibility complex II (MHCII)-APC-Cy7 (clone: M5/114.15.2; Biolegend, #107627, diluted 1:200). .. All samples were fixed in 1 % PFA, and Trucount counting beads (5000 beads/sample; BD Trucount Absolute Counting Tubes, BD Biosciences, #663028) were added for use in estimation of total cell numbers.
Immunopeptidomics:Article Title: Systemic inhibition of soluble TNF significantly changes glial cell populations leading to improved myelin integrity and better functional outcome after experimental stroke
Article Snippet: .. Glial cell samples were incubated with CD45-PerCP-Cy 5.5 (clone: 30-F11; BD Biosciences, #561869, diluted 1:100), astrocyte cell surface antigen-2 (ACSA-2)-FITC (clone: REA969; Miltenyi Biotec, #130–116–243, diluted 1:50), CD140a-APC (PDGFRα; clone: REA637; Miltenyi Biotec, #130–122–942, diluted 1:10), and O4-PE (clone: REA576; Miltenyi Biotec, #130–117–823, diluted 1:50), while immune cell samples were incubated with CD45-PerCP-Cy 5.5 (clone: 30-F11), CD11b-BB515 (clone: M1/70; BD Biosciences, #564455, diluted 1:200), CD4-APC (clone: RM4–5; BD Biosciences, #561091, diluted 1:200), CD8a-PE (clone: 53–6.7; BD Biosciences #553032, diluted 1:200), Lymphocyte antigen 6 G (Ly-6G)-BV786 (clone: 1A8; BD Biosciences, #740953, diluted 1:200), and Major histocompatibility complex II (MHCII)-APC-Cy7 (clone: M5/114.15.2; Biolegend, #107627, diluted 1:200). .. All samples were fixed in 1 % PFA, and Trucount counting beads (5000 beads/sample; BD Trucount Absolute Counting Tubes, BD Biosciences, #663028) were added for use in estimation of total cell numbers.
Article Title: Systemic inhibition of soluble TNF significantly changes glial cell populations leading to improved myelin integrity and better functional outcome after experimental stroke.
Article Snippet: .. Glial cell samples were incubated with CD45-PerCP-Cy5.5 (clone: 30-F11; BD Biosciences, #561869, diluted 1:100), astrocyte cell surface antigen-2 (ACSA-2)-FITC (clone: REA969; Miltenyi Biotec, #130–116–243, diluted 1:50), CD140a-APC (PDGFRα; clone: REA637; Miltenyi Biotec, #130–122–942, diluted 1:10), and O4-PE (clone: REA576; Miltenyi Biotec, #130–117–823, diluted 1:50), while immune cell samples were incubated with CD45-PerCP-Cy5.5 (clone: 30-F11), CD11b-BB515 (clone: M1/70; BD Biosciences, #564455, diluted 1:200), CD4-APC (clone: RM4–5; BD Biosciences, #561091, diluted 1:200), CD8a-PE (clone: 53–6.7; BD Biosciences #553032, diluted 1:200), Lymphocyte antigen 6 G (Ly-6G)-BV786 (clone: 1A8; BD Biosciences, #740953, diluted 1:200), and Major histocompatibility complex II (MHCII)-APC-Cy7 (clone: M5/114.15.2; Biolegend, #107627, diluted 1:200). .. All samples were fixed in 1 % PFA, and Trucount counting beads (5000 beads/sample; BD Trucount Absolute Counting Tubes, BD Biosciences, #663028) were added for use in estimation of total cell numbers.
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