Journal: Cell Death & Disease

Article Title: The protective effects of systemic dexamethasone on sensory epithelial damage and hearing loss in targeted Cx26-null mice

doi: 10.1038/s41419-022-04987-3

Figure Lengend Snippet: A Schematic of the experimental workflow, Cx26 f/f ; Fgfr3-CreER mice were treated with TMX and then treated with dexamethasone 5 mg /kg of body weight for five days. B – E Immunofluorescence staining with DAPI to show the pattern of hair cell damage from different group. F – I Immunofluorescence staining with CD45 (green) to visualized the distribution and morphology of macrophages in the basilar membrane from different groups. J – M The three-dimensional reconstruction of CD45 positive cells to show the morphology of CD45 positive cells from different groups. N Quantification of the numbers of CD45 positive cells on the scala tympani side of the basilar membrane in different groups. O Quantifications of CD45 positive cells size in three turns from different group. n = 4 mice in each group, ** P < 0.01, *** P < 0.001. Scale in panels ( B , C ) represents for 40 μm.

Article Snippet: After being rinsed three times with 0.1% Tween-20 in PBS (PBST), the flattened cochlear preparations or cochlear explants were incubated in a blocking solution of 5% Bovine Serum Albumin, followed by incubation with primary antibodies: polyclonal rabbit anti-myosin7a antibodies (25-6790, Proteus Bio-Sciences), polyclonal goat anti-sox2 antibodies (sc-17320, Santa Cruz Biotechnology), polyclonal rabbit anti-Cx26 antibodies (512800, Invitrogen) and goat anti-CD45 polyclonal antibody (AF114, R&D Systems) diluted in PBS overnight at 4 °C.

Techniques: Immunofluorescence, Staining, Membrane

Journal: Laboratory investigation; a journal of technical methods and pathology

Article Title: Characterization of immune cell subtypes in three commonly used mouse strains reveals gender and strain-specific variations

doi: 10.1038/s41374-018-0137-1

Figure Lengend Snippet: Antibody combinations used in immunophenotyping

Article Snippet: Fixation/Permeabilization Solution Kit with BD Golgi-PlugTM kit and Mouse Foxp3 Buffer Set was used for intracellular staining and purchased from BD Biosciences. table ft1 table-wrap mode="anchored" t5 caption a7 iMC CD11b Vio Blue Gr-1 APC-eFluor780 * Ly6b FITC Macrophages CD11b Vio Blue F4/80 APC CD68 PE mDC CD11b Vio Blue CD11c PE CD80 APC CD86 FITC pDC * CD11b Vio Blue CD11c PE B220 Vio770 Siglec H FITC CD4 T-cells CD3 APC-Cy7 CD4 eFluor450 CD4 T-regs CD3 APC-Cy7 CD4 eFluor450 CD25 PE FoxP3APC CD8 T-cells CD3 APC-Cy7 CD8 FITC B cells B220 Vio770 CD19 PE NK cells CD3 APC-Cy7 NKp46 FITC Neutrophils CD11b Vio Blue Gr-1 APC-eFluor780 Ly6b FITC * F4/80 APC Open in a separate window * negative tor this antibody Antibody combinations used in immunophenotyping table ft1 table-wrap mode="anchored" t5 caption a7 Antibody Fluorochrome Vendor Clone Catalogue # Antibody Concentration (in 100 pL FACS buffer) CD3e APC-Cy7 Fisher Scientific 145–2C11 BDB557596 1 pL CD4 eFluor450 eBioscience GK1.5 48–0041-82 1 pL CD8 FITC eBioscience 53–6.7 11–0081-82 0.6 pL CD11b Vio Blue Miltenyi M1/70.15.11.5 130–097-336 3.75 pL CD11c PE Miltenyi N418 130102545 3.75 pL CD19 PE Miltenyi 6D5 130–102-598 5 pL CD25 PE eBioscience PC61.5 12–0251-81 0.5 pL CD68 PE Miltenyi FA-11 130–102-614 3.75 pL CD80 APC Miltenyi 16–10A1 130–102-584 3.75 pL CD86 FITC Miltenyi PO3.3 130–102-506 5 pL B220 APC-Vio770 Miltenyi RA3–6B2 130–102-267 3.75 pL F4/80 APC Miltenyi REA126 130–102-379 3.75 pL FoxP3 APC eBioscience FJK-16s 17–5773-82 5 pL Gr1 APC-eFluor780 eBioscience RB6–8C5 47–5931-80 1.2 pL Ly6b FITC abcam 7/4 ab53453 0.65 pL NKp46 FITC Miltenyi 29A1.4.9 130–102-300 3.75 pL SiglecH FITC eBioscience eBio440c 11–0333-82 0.5 pL Open in a separate window Details of antibodies used in the study

Techniques:

Journal: Laboratory investigation; a journal of technical methods and pathology

Article Title: Characterization of immune cell subtypes in three commonly used mouse strains reveals gender and strain-specific variations

doi: 10.1038/s41374-018-0137-1

Figure Lengend Snippet: Details of antibodies used in the study

Article Snippet: Fixation/Permeabilization Solution Kit with BD Golgi-PlugTM kit and Mouse Foxp3 Buffer Set was used for intracellular staining and purchased from BD Biosciences. table ft1 table-wrap mode="anchored" t5 caption a7 iMC CD11b Vio Blue Gr-1 APC-eFluor780 * Ly6b FITC Macrophages CD11b Vio Blue F4/80 APC CD68 PE mDC CD11b Vio Blue CD11c PE CD80 APC CD86 FITC pDC * CD11b Vio Blue CD11c PE B220 Vio770 Siglec H FITC CD4 T-cells CD3 APC-Cy7 CD4 eFluor450 CD4 T-regs CD3 APC-Cy7 CD4 eFluor450 CD25 PE FoxP3APC CD8 T-cells CD3 APC-Cy7 CD8 FITC B cells B220 Vio770 CD19 PE NK cells CD3 APC-Cy7 NKp46 FITC Neutrophils CD11b Vio Blue Gr-1 APC-eFluor780 Ly6b FITC * F4/80 APC Open in a separate window * negative tor this antibody Antibody combinations used in immunophenotyping table ft1 table-wrap mode="anchored" t5 caption a7 Antibody Fluorochrome Vendor Clone Catalogue # Antibody Concentration (in 100 pL FACS buffer) CD3e APC-Cy7 Fisher Scientific 145–2C11 BDB557596 1 pL CD4 eFluor450 eBioscience GK1.5 48–0041-82 1 pL CD8 FITC eBioscience 53–6.7 11–0081-82 0.6 pL CD11b Vio Blue Miltenyi M1/70.15.11.5 130–097-336 3.75 pL CD11c PE Miltenyi N418 130102545 3.75 pL CD19 PE Miltenyi 6D5 130–102-598 5 pL CD25 PE eBioscience PC61.5 12–0251-81 0.5 pL CD68 PE Miltenyi FA-11 130–102-614 3.75 pL CD80 APC Miltenyi 16–10A1 130–102-584 3.75 pL CD86 FITC Miltenyi PO3.3 130–102-506 5 pL B220 APC-Vio770 Miltenyi RA3–6B2 130–102-267 3.75 pL F4/80 APC Miltenyi REA126 130–102-379 3.75 pL FoxP3 APC eBioscience FJK-16s 17–5773-82 5 pL Gr1 APC-eFluor780 eBioscience RB6–8C5 47–5931-80 1.2 pL Ly6b FITC abcam 7/4 ab53453 0.65 pL NKp46 FITC Miltenyi 29A1.4.9 130–102-300 3.75 pL SiglecH FITC eBioscience eBio440c 11–0333-82 0.5 pL Open in a separate window Details of antibodies used in the study

Techniques: Concentration Assay

Journal: Laboratory investigation; a journal of technical methods and pathology

Article Title: Characterization of immune cell subtypes in three commonly used mouse strains reveals gender and strain-specific variations

doi: 10.1038/s41374-018-0137-1

Figure Lengend Snippet: Cell suspensions from BM and spleen were isolated from 8-week old mice. The cells were stained with B220 and CD19 antibodies for B cells, and CD3 and NKp46 antibodies for NK cells. The cells were then subjected to flow cytometry to determine cell-type percentages. (A) B cells in BM, (n≥10). (B) B cells in the spleen, (n ≥11). (C) NK cells in BM, (n≥9). (D) NK cells in the spleen, (n≥7). Results are shown as scatter plots depicting average cell percentages (percent of live cells). Error bars denote SEM. Each dot represents the value from a single mouse. (♂) and (♀) represent male and female mice respectively. **p ≤ 0.01, ***p ≤ 0.001, ****p≤0.0001.

Article Snippet: Fixation/Permeabilization Solution Kit with BD Golgi-PlugTM kit and Mouse Foxp3 Buffer Set was used for intracellular staining and purchased from BD Biosciences. table ft1 table-wrap mode="anchored" t5 caption a7 iMC CD11b Vio Blue Gr-1 APC-eFluor780 * Ly6b FITC Macrophages CD11b Vio Blue F4/80 APC CD68 PE mDC CD11b Vio Blue CD11c PE CD80 APC CD86 FITC pDC * CD11b Vio Blue CD11c PE B220 Vio770 Siglec H FITC CD4 T-cells CD3 APC-Cy7 CD4 eFluor450 CD4 T-regs CD3 APC-Cy7 CD4 eFluor450 CD25 PE FoxP3APC CD8 T-cells CD3 APC-Cy7 CD8 FITC B cells B220 Vio770 CD19 PE NK cells CD3 APC-Cy7 NKp46 FITC Neutrophils CD11b Vio Blue Gr-1 APC-eFluor780 Ly6b FITC * F4/80 APC Open in a separate window * negative tor this antibody Antibody combinations used in immunophenotyping table ft1 table-wrap mode="anchored" t5 caption a7 Antibody Fluorochrome Vendor Clone Catalogue # Antibody Concentration (in 100 pL FACS buffer) CD3e APC-Cy7 Fisher Scientific 145–2C11 BDB557596 1 pL CD4 eFluor450 eBioscience GK1.5 48–0041-82 1 pL CD8 FITC eBioscience 53–6.7 11–0081-82 0.6 pL CD11b Vio Blue Miltenyi M1/70.15.11.5 130–097-336 3.75 pL CD11c PE Miltenyi N418 130102545 3.75 pL CD19 PE Miltenyi 6D5 130–102-598 5 pL CD25 PE eBioscience PC61.5 12–0251-81 0.5 pL CD68 PE Miltenyi FA-11 130–102-614 3.75 pL CD80 APC Miltenyi 16–10A1 130–102-584 3.75 pL CD86 FITC Miltenyi PO3.3 130–102-506 5 pL B220 APC-Vio770 Miltenyi RA3–6B2 130–102-267 3.75 pL F4/80 APC Miltenyi REA126 130–102-379 3.75 pL FoxP3 APC eBioscience FJK-16s 17–5773-82 5 pL Gr1 APC-eFluor780 eBioscience RB6–8C5 47–5931-80 1.2 pL Ly6b FITC abcam 7/4 ab53453 0.65 pL NKp46 FITC Miltenyi 29A1.4.9 130–102-300 3.75 pL SiglecH FITC eBioscience eBio440c 11–0333-82 0.5 pL Open in a separate window Details of antibodies used in the study

Techniques: Isolation, Staining, Flow Cytometry

Journal: Laboratory investigation; a journal of technical methods and pathology

Article Title: Characterization of immune cell subtypes in three commonly used mouse strains reveals gender and strain-specific variations

doi: 10.1038/s41374-018-0137-1

Figure Lengend Snippet: Cell suspensions from BM and spleen were isolated from 8-week old mice. The cells were stained with CD11c, B220 and Siglec H antibodies. CD11b antibody was included as a negative marker for pDCs. The cells were then subjected to flow cytometry to determine cell-type percentages. (A) pDCs in BM, (n≥11). (B) pDCs in the spleen, (n ≥9). Results are shown as scatter plots depicting average cell percentages (percent of live cells). Error bars denote SEM. Each dot represents the value from a single mouse. (♂) and (♀) represent male and female mice respectively.*p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001, ****p≤0.0001.

Article Snippet: Fixation/Permeabilization Solution Kit with BD Golgi-PlugTM kit and Mouse Foxp3 Buffer Set was used for intracellular staining and purchased from BD Biosciences. table ft1 table-wrap mode="anchored" t5 caption a7 iMC CD11b Vio Blue Gr-1 APC-eFluor780 * Ly6b FITC Macrophages CD11b Vio Blue F4/80 APC CD68 PE mDC CD11b Vio Blue CD11c PE CD80 APC CD86 FITC pDC * CD11b Vio Blue CD11c PE B220 Vio770 Siglec H FITC CD4 T-cells CD3 APC-Cy7 CD4 eFluor450 CD4 T-regs CD3 APC-Cy7 CD4 eFluor450 CD25 PE FoxP3APC CD8 T-cells CD3 APC-Cy7 CD8 FITC B cells B220 Vio770 CD19 PE NK cells CD3 APC-Cy7 NKp46 FITC Neutrophils CD11b Vio Blue Gr-1 APC-eFluor780 Ly6b FITC * F4/80 APC Open in a separate window * negative tor this antibody Antibody combinations used in immunophenotyping table ft1 table-wrap mode="anchored" t5 caption a7 Antibody Fluorochrome Vendor Clone Catalogue # Antibody Concentration (in 100 pL FACS buffer) CD3e APC-Cy7 Fisher Scientific 145–2C11 BDB557596 1 pL CD4 eFluor450 eBioscience GK1.5 48–0041-82 1 pL CD8 FITC eBioscience 53–6.7 11–0081-82 0.6 pL CD11b Vio Blue Miltenyi M1/70.15.11.5 130–097-336 3.75 pL CD11c PE Miltenyi N418 130102545 3.75 pL CD19 PE Miltenyi 6D5 130–102-598 5 pL CD25 PE eBioscience PC61.5 12–0251-81 0.5 pL CD68 PE Miltenyi FA-11 130–102-614 3.75 pL CD80 APC Miltenyi 16–10A1 130–102-584 3.75 pL CD86 FITC Miltenyi PO3.3 130–102-506 5 pL B220 APC-Vio770 Miltenyi RA3–6B2 130–102-267 3.75 pL F4/80 APC Miltenyi REA126 130–102-379 3.75 pL FoxP3 APC eBioscience FJK-16s 17–5773-82 5 pL Gr1 APC-eFluor780 eBioscience RB6–8C5 47–5931-80 1.2 pL Ly6b FITC abcam 7/4 ab53453 0.65 pL NKp46 FITC Miltenyi 29A1.4.9 130–102-300 3.75 pL SiglecH FITC eBioscience eBio440c 11–0333-82 0.5 pL Open in a separate window Details of antibodies used in the study

Techniques: Isolation, Staining, Marker, Flow Cytometry