Journal: Physiological Reports

Article Title: Autologous minced muscle grafts improve endogenous fracture healing and muscle strength after musculoskeletal trauma

doi: 10.14814/phy2.13362

Figure Lengend Snippet: Flow cyotometry antibodies

Article Snippet: CD68 , ED1 , Novus Biologicals , NB600‐985V , 0.001 mg/mL.

Techniques: Concentration Assay

Journal: Journal of vascular research

Article Title: A Novel Model of Balloon Angioplasty Injury in Rat Arteriovenous Fistula

doi: 10.1159/000507080

Figure Lengend Snippet: (A) α-SMA and CD68 immunostaining of representative sections from Group 1 and Group 2. Scale bar=100 μM. Higher magnification images are at 40X. (B) Quantification of α-SMA (intimal region) and CD68 (intimal and medial regions) expression from AVF-artery and AVF-vein. N=3–4 in each group. *p<0.05, **p<0.001. An unpaired t-test was used to test for statistical differences between AVF without angioplasty and AVF with angioplasty groups.

Article Snippet: Immunohistochemical Analysis Paraffin embedded arterial and venous sections were evaluated to identify the cells contributing to the vascular remodeling following balloon angioplasty. α-SMA (14-9760-82, Invitrogen) and CD68 (NB600–985, Novus Biologicals) primary antibodies were used to verify the presence of smooth muscle cell phenotype and inflammatory cells respectively. α-SMA expression in the intimal region and CD68 expression in both medial and intimal regions of AVF vessels were quantified using Cellsense Dimension Software (Olympus Life Science) on digital photographs taken at a final magnification of 20X.

Techniques: Immunostaining, Expressing

Journal: Frontiers in Immunology

Article Title: Site-Specific Reprogramming of Macrophage Responsiveness to Bacterial Lipopolysaccharide in Obesity

doi: 10.3389/fimmu.2019.01496

Figure Lengend Snippet: Macrophage abundance and cytokine secretion in AT-SVF cultures. (A,B) Representative flow-cytometry density plots of staining for CD45 (a leukocyte marker) and CD68 (a rat macrophage marker) in AT-SVF cells from LFD-fed rats (A) or HFD-fed rats (B) . Stained cells are shown in red; unstained cells are shown in gray. Group results— n = 5 culture replicates in (A) ; n = 12 in (B) —are shown as 95% confidence intervals within each quadrant. (C) Representative histograms showing that LPS-stimulated cells that are CD45 + /CD68 + (macrophages) stain more strongly for TNF-α than the other cellular phenotypes present in the AT-SVF culture. For staining intensities of all samples, see . (D–F) Effects of diet and ex-vivo stimulation with LPS on the secretion of TNF-α (D) , IL-1β (E) and IL-10 (F) by AT-SVF cells. To aid comparison of this macrophage subpopulation with the others, the scales in these panels are identical to those of the corresponding panels in , . Data in (D–F) are expressed as means ± SEM. Sample size (number of culture replicates) was 4 in the LFD group not stimulated with LPS (saline), 6 in the HFD group not stimulated with LPS (saline), 5 in the LFD group stimulated with LPS, and 7 in the HFD group stimulated with LPS. (G) TNF-α/IL-10 and IL-1β/IL-10 ratios (means ± SEM) in the LPS-stimulated AT-SVF cultures from the LFD vs. HFD groups. Statistical marks: a significant effect of LPS (compared to saline), b significant effect of diet.

Article Snippet: Stainings were achieved by 30-min incubation with the following monoclonal antibodies at 4°C and in the dark: mouse anti-rat CD45 conjugated with APC-Cy7 (BD 561586; 1:100); mouse anti-rat CD68 conjugated with PE (Novus Biologicals NB600-985PE; 1:100); and hamster anti-rat/mouse TNF-α conjugated with APC (BioLegend 506107; 1:25).

Techniques: Flow Cytometry, Staining, Marker, Ex Vivo, Comparison, Saline

Journal: Frontiers in Immunology

Article Title: Site-Specific Reprogramming of Macrophage Responsiveness to Bacterial Lipopolysaccharide in Obesity

doi: 10.3389/fimmu.2019.01496

Figure Lengend Snippet: Macrophage abundance and cytokine secretion in alveolar cultures. (A,B) Representative flow-cytometry density plots of staining for CD45 (a leukocyte marker) and CD68 (a rat macrophage marker) in alveolar cells from LFD-fed rats (A) or HFD-fed rats (B) . Stained cells are shown in red; unstained cells are shown in gray. Group results— n = 5 culture replicates in (A) ; n = 8 in (B) —are shown as 95% confidence intervals within each quadrant. (C) Representative histograms showing that LPS-stimulated cells that are CD45 + /CD68 + (macrophages) stain strongly for TNF-α. Because CD45 + /CD68 − were very rare in the alveolar culture, their histograms could not be reliably determined and, consequently, this subpopulation is not shown in (C) of this particular figure. For staining intensities of all samples, see . (D–F) Effects of diet and ex-vivo stimulation with LPS on the secretion of TNF-α (D) , IL-1β (E) , and IL-10 (F) by alveolar macrophages. To aid comparison of this macrophage subpopulation with the others, the scales in these panels are identical to those of the corresponding panels in , . Data in (D–F) are expressed as means ± SEM. Sample size (number of culture replicates) was 6 in the LFD group not stimulated with LPS (saline), 7 in the HFD group not stimulated with LPS (saline), 18 in the LFD group stimulated with LPS, and 16 in the HFD group stimulated with LPS. (G) TNF-α/IL-10 and IL-1β/IL-10 ratios (means ± SEM) in the LPS-stimulated alveolar macrophages from the LFD vs. HFD groups. Statistical mark: a significant effect of LPS (compared to saline).

Article Snippet: Stainings were achieved by 30-min incubation with the following monoclonal antibodies at 4°C and in the dark: mouse anti-rat CD45 conjugated with APC-Cy7 (BD 561586; 1:100); mouse anti-rat CD68 conjugated with PE (Novus Biologicals NB600-985PE; 1:100); and hamster anti-rat/mouse TNF-α conjugated with APC (BioLegend 506107; 1:25).

Techniques: Flow Cytometry, Staining, Marker, Ex Vivo, Comparison, Saline

Journal: Frontiers in Immunology

Article Title: Site-Specific Reprogramming of Macrophage Responsiveness to Bacterial Lipopolysaccharide in Obesity

doi: 10.3389/fimmu.2019.01496

Figure Lengend Snippet: Macrophage abundance and cytokine secretion in peritoneal cultures. (A,B) Representative flow-cytometry density plots of staining for CD45 (a leukocyte marker) and CD68 (a rat macrophage marker) in peritoneal cells from LFD-fed rats (A) or HFD-fed rats (B) . Stained cells are shown in red; unstained cells are shown in gray. Group results— n = 5 culture replicates in (A) ; n = 8 in (B) —are shown as 95% confidence intervals within each quadrant. (C) Representative histograms showing that LPS-stimulated cells that are CD45 + /CD68 + (macrophages) stain more strongly for TNF-α than the other cellular phenotypes present in the peritoneal culture. For staining intensities of all samples, see . (D–F) Effects of diet and ex-vivo stimulation with LPS on the secretion of TNF-α (D) , IL-1β (E) , and IL-10 (F) by peritoneal macrophages. To aid comparison of this macrophage subpopulation with the others, the scales in these panels are identical to those of the corresponding panels in , . Data are expressed as means ± SEM. Sample size (number of culture replicates) was 12 in the LFD group not stimulated with LPS (saline), 7 in the HFD group not stimulated with LPS (saline), 16 in the LFD group stimulated with LPS, and 13 in the HFD group stimulated with LPS. (G) TNF-α/IL-10 and IL-1β/IL-10 ratios (means ± SEM) in the LPS-stimulated peritoneal macrophages from the LFD vs. HFD groups. Statistical marks: a significant effect of LPS (compared to saline), b significant effect of diet.

Article Snippet: Stainings were achieved by 30-min incubation with the following monoclonal antibodies at 4°C and in the dark: mouse anti-rat CD45 conjugated with APC-Cy7 (BD 561586; 1:100); mouse anti-rat CD68 conjugated with PE (Novus Biologicals NB600-985PE; 1:100); and hamster anti-rat/mouse TNF-α conjugated with APC (BioLegend 506107; 1:25).

Techniques: Flow Cytometry, Staining, Marker, Ex Vivo, Comparison, Saline

Journal: iScience

Article Title: A rat liver cell atlas reveals intrahepatic myeloid heterogeneity

doi: 10.1016/j.isci.2023.108213

Figure Lengend Snippet:

Article Snippet: Mouse anti-rat CD68 AF700; Clone: ED1 , Novus Biologicals , Cat# NB600-985AF700.

Techniques: Recombinant, SYBR Green Assay, Staining, Single Cell, Selection, Gene Expression, Software

Journal: Science Advances

Article Title: A nanofiber-hydrogel composite improves tissue repair in a rat model of Crohn’s disease perianal fistulas

doi: 10.1126/sciadv.ade1067

Figure Lengend Snippet: ( A ) Modeling and treatment timeline of CD-PAF in a rat model. ( B ) Steps in rat surgery to induce fistulas and then treat them with mfNHC. ( C ) MRI images at day 28 demonstrating patent fistula tracts (axial T1 images were obtained with setons in place, and coronal T1 images were obtained after setons were cut). ( D ) MRI images showing inflammation around fistula tracts (left, coronal T1, yellow arrow) and occasional abscess formation at day 28 (right, axial T2, yellow arrow). ( E ) Follow-up MRI at day 42 showing patent fistulas (yellow arrow) 14 days after removing setons at day 28. Note that there was no healing and spontaneous closing. ( F ) H&E staining of the fistula tract with a partially re-epithelialized lumen surrounded by dense acute and chronic inflammation and peri-fistula abscess formation (left scale bar, 2.5 mm; middle scale bar, 1 mm; right scale bar, 250 μm). ( G ) IF staining demonstrating PANCK + epithelial cells lining fistula lumens. Green, PANCK; blue, DAPI (scale bar, 200 μm). ( H to J ) IF staining demonstrating the spatial arrangement of the inflammatory milieu around the fistula tracts. MPO + , neutrophils; CD68 + CD163 + , macrophages; CD20 + , B cells; CD45RO + , memory T lymphocytes (scale bar, 500 μm).

Article Snippet: The following reagents were used in the studies: anti-CD68 mouse antibody (ab31630, Abcam, Cambridge, UK), anti-CD68 rabbit antibody (ab125212, Abcam, Cambridge, UK), Alexa Flour 647 anti-rat CD68/SR-D1 (Novus Biologicals, Littleton, CO), anti-CD86 mouse antibody (ab213044, Abcam, Cambridge, UK), anti-CD38 antibody (GTX37752, GeneTex, Irvine, CA), recombinant anti-CD163 rabbit antibody (ab182422, Abcam, Cambridge, UK), phycoerthrin (PE)/Cy5.5 anti-rat CD163 (Novus Biologicals, Littleton, CO), PE anti-rat CD38 (BioLegend, San Diego, CA), anti–α-SMA rabbit antibody (ab5694, Abcam, Cambridge, UK), mouse CD45RO antibody (MA5-11532, Thermo Fisher Scientific, Waltham, MA), anti-CD20 antibody (70168, Cell Signaling Technology, Danvers, MA), Cytokeratin Pan Type I/II Antibody Cocktail mouse (MA1-82041, Thermo Fisher Scientific, Waltham, MA), Alexa Fluor 594 or 488 and 647 secondary antibodies (Life Technologies, Carlsbad, CA), Rat BD Fc Block (550270, BD Biosciences, San Jose, CA), BV421 mouse anti-rat CD3 (563948, BD Biosciences, San Jose, CA), peridin chlorophyll protein (PerCP)/Cyanine5.5 anti-rat CD11b/c antibody (201819, BioLegend, San Diego, CA), PE/Cyanine7 anti-rat CD45 antibody (202213, BioLegend, San Diego, CA), CD68/SR-D1 antibody (ED1) [Alexa Fluor 405] (NB600-985/AF405, Novus Biologicals, Littleton, CO), PE mouse anti-rat CD86 (551396, BD Biosciences, San Jose, CA), PE/Cyanine7 anti-rat CD161 antibody (205609, BioLegend, San Diego, CA), PE/Cyanine7 anti-rat CD161 antibody (202307, BioLegend, San Diego, CA), fluorescein isothiocyanate (FITC) anti-mouse/rat CD29 antibody (102205, BioLegend, San Diego, CA), PE mouse anti-rat CD4 (551397, BD Biosciences, San Jose, CA), allophycocyanin (APC) mouse anti-rat CD90/mouse CD90.1 (561409, BD Biosciences, San Jose, CA), PE anti-rat CD106 antibody (200403, BioLegend, San Diego, CA), CD31 monoclonal antibody (TLD-3A12), FITC (MA5-16952, Thermo Fisher Scientific, Waltham, MA), FITC anti-rat CD45 antibody (202205, BioLegend, San Diego, CA), rat IL-6 DuoSet ELISA kit (DY522-05, R&D Systems, Minneapolis, MN), rat IL-10 DuoSet ELISA kit (DY506-05, R&D Systems, Minneapolis, MN), and rat TNFα ELISA kit (ab100785, Abcam, Cambridge, UK).

Techniques: Staining

Journal: Science Advances

Article Title: A nanofiber-hydrogel composite improves tissue repair in a rat model of Crohn’s disease perianal fistulas

doi: 10.1126/sciadv.ade1067

Figure Lengend Snippet: ( A ) Staining of MPO + neutrophils and CD45RO + T cells. ( B ) Staining of CD20 + B cells and CD45RO + memory T cells. ( C ) Staining of CD68 + macrophages and CD163 + M2-like macrophages (top scale bar, 200 μm; bottom scale bar, 100 μm).

Article Snippet: The following reagents were used in the studies: anti-CD68 mouse antibody (ab31630, Abcam, Cambridge, UK), anti-CD68 rabbit antibody (ab125212, Abcam, Cambridge, UK), Alexa Flour 647 anti-rat CD68/SR-D1 (Novus Biologicals, Littleton, CO), anti-CD86 mouse antibody (ab213044, Abcam, Cambridge, UK), anti-CD38 antibody (GTX37752, GeneTex, Irvine, CA), recombinant anti-CD163 rabbit antibody (ab182422, Abcam, Cambridge, UK), phycoerthrin (PE)/Cy5.5 anti-rat CD163 (Novus Biologicals, Littleton, CO), PE anti-rat CD38 (BioLegend, San Diego, CA), anti–α-SMA rabbit antibody (ab5694, Abcam, Cambridge, UK), mouse CD45RO antibody (MA5-11532, Thermo Fisher Scientific, Waltham, MA), anti-CD20 antibody (70168, Cell Signaling Technology, Danvers, MA), Cytokeratin Pan Type I/II Antibody Cocktail mouse (MA1-82041, Thermo Fisher Scientific, Waltham, MA), Alexa Fluor 594 or 488 and 647 secondary antibodies (Life Technologies, Carlsbad, CA), Rat BD Fc Block (550270, BD Biosciences, San Jose, CA), BV421 mouse anti-rat CD3 (563948, BD Biosciences, San Jose, CA), peridin chlorophyll protein (PerCP)/Cyanine5.5 anti-rat CD11b/c antibody (201819, BioLegend, San Diego, CA), PE/Cyanine7 anti-rat CD45 antibody (202213, BioLegend, San Diego, CA), CD68/SR-D1 antibody (ED1) [Alexa Fluor 405] (NB600-985/AF405, Novus Biologicals, Littleton, CO), PE mouse anti-rat CD86 (551396, BD Biosciences, San Jose, CA), PE/Cyanine7 anti-rat CD161 antibody (205609, BioLegend, San Diego, CA), PE/Cyanine7 anti-rat CD161 antibody (202307, BioLegend, San Diego, CA), fluorescein isothiocyanate (FITC) anti-mouse/rat CD29 antibody (102205, BioLegend, San Diego, CA), PE mouse anti-rat CD4 (551397, BD Biosciences, San Jose, CA), allophycocyanin (APC) mouse anti-rat CD90/mouse CD90.1 (561409, BD Biosciences, San Jose, CA), PE anti-rat CD106 antibody (200403, BioLegend, San Diego, CA), CD31 monoclonal antibody (TLD-3A12), FITC (MA5-16952, Thermo Fisher Scientific, Waltham, MA), FITC anti-rat CD45 antibody (202205, BioLegend, San Diego, CA), rat IL-6 DuoSet ELISA kit (DY522-05, R&D Systems, Minneapolis, MN), rat IL-10 DuoSet ELISA kit (DY506-05, R&D Systems, Minneapolis, MN), and rat TNFα ELISA kit (ab100785, Abcam, Cambridge, UK).

Techniques: Staining