cd11b percp cyanine5 5  (Elabscience Biotechnology)

Journal: bioRxiv

Article Title: Spinal Cord Microglia Exhibit Impaired Repair Responses to Myelin Damage

doi: 10.64898/2026.02.23.707565

Figure Lengend Snippet: (A). Flow cytometry workflow. The total leukocyte population was gated away from debris. (B) Single cells were then gated away from any multiplets. (C) Viable cells expressing low levels of CD45 were gated away from other cells and infiltrating macrophages. (D) Lastly, cells expressing low levels of CD45 and positive for CD11b were selected for the analysis, as they are microglia. (E) Microglia as a percentage of the total single cells in the sample graphed to highlight the comparison between brain and spinal cord. Data represents mean and error bars represent standard error of the mean. Statistical analysis was performed using a 3-way ANOVA with Tukey’s post-hoc analysis, n = 7 – 10 where each sample represents a different animal, * p < 0.05, ** p < 0.01, *** p < 0.001, **** p <0.0001.

Article Snippet: The resulting cells were incubated in an FC block solution (Tonbo 70-0161, 1:100) for 10 minutes and then stained in an antibody mix for 30 minutes which consisted of CD45 violetFluor 450 (Tonbo 75-0451, 1:75), CD11b PerCP-Cyanine5.5 (Tonbo 65-0112, 1:75), Brilliant Violet 605 anti-mouse CD16 (Biolegend 158027, 1:75), FITC anti-mouse CD68 (Biolegend 137006, 1:75), APC/Fire 810 anti-mouse CX3CR1 (Biolegend 149054, 1:75), APC anti-mouse F4/80 (Biolegend 123116, 1:75), APC/Fire 750 anti-mouse CD73 (Biolegend 127221, 1:75), PE anti-mouse TREM2 (R&D Systems FAB17291P, 1:75), and viability dye (Tonbo 13-0871, 1:1000).

Techniques: Flow Cytometry, Expressing, Comparison

Journal: Advanced Science

Article Title: Rescuing Mitochondrial Dysfunction in Macrophages Prevents Osteonecrosis of the Jaw in Anti‐Resorptive Therapy

doi: 10.1002/advs.202517586

Figure Lengend Snippet: ZA promotes macrophage pro‐inflammatory polarization and induces BRONJ. A) Representative H&E‐stained images of tooth extraction site (TES). Red dashed lines indicate osteonecrosis area (Nec). Scale bars: 200 µm. B) Micro‐CT analysis of new bone formation in TES. C) The panel shows the quantitative data of open wound area ( n = 5). D) Immunofluorescence (IF) imaging of CD31 hi EMCN hi type‐H vessels in tooth extraction socket (TES). Scale bars: 100 µm. E) Gating strategy and flow cytometry assessment of jawbone macrophage stained with CD45, CD11b, F4/80, CD86, and CD206 antibodies. F) IF assay of F4/80 + CD86 + or F4/80 + CD206 + macrophages in TES. White arrows indicate representative cells. Scale bars: 100 µm. G) Western blot revealing the total protein expression of iNOS, CD206, pro IL‐1β, and IL‐1β in BMDMs treated with ZA, LPS, or IL‐4 for 24 h. H) Schematic depicting the bulk RNA sequencing process for Ctrl or ZA‐treated BMDMs. I) Volcano plot of gene expression levels determined by bulk RNA sequencing. Deep green and yellow plots represent genes that were significantly differentially expressed. J) Top pathways identified by GO enrichment analysis from differentially expressed genes of BMDMs after ZA treatment. Results are representative of at least three independent experiments performed. Densitometric quantifications are indicated beneath the representative blots. Panels show the quantitative data of osteonecrosis area (A, n = 5), trabecular bone volume/tissue volume (BV/TV), and bone mineral density (BMD) in TES (B, n = 5), type‐H vessel fraction (D, n = 5), and M1‐like and M2‐like macrophage numbers (F, n = 5). Statistical significance was examined by two‐tailed Student's t ‐tests or a two‐sided unpaired Welch's t ‐tests (A–D, F). Results are presented as the mean ± SD.

Article Snippet: [ ] The cells were washed with PBS and stained with Fixable Viability Dye eFluor 506 (Invitrogen, 65‐0866‐14), APC‐eFlour 780 Anti‐Mouse CD45 (Invitrogen, 47‐0451‐82), PerCP/Cyanine5.5 Anti‐Mouse CD11b (Elabscience, E‐AB‐F1081J), Brilliant Violet 605 Anti‐Mouse F4/80 (Biolegend, 123133), PE Anti‐Mouse CD86 (BD Biosciences, 561963), and FITC Anti‐Mouse CD206 (Biolegend, 141703) antibodies.

Techniques: Staining, Extraction, Micro-CT, Immunofluorescence, Imaging, Flow Cytometry, Western Blot, Expressing, RNA Sequencing, Gene Expression, Two Tailed Test

Journal: Advanced Science

Article Title: Rescuing Mitochondrial Dysfunction in Macrophages Prevents Osteonecrosis of the Jaw in Anti‐Resorptive Therapy

doi: 10.1002/advs.202517586

Figure Lengend Snippet: Deficiency of autophagosome maturation in Lyz2cre; Atg5 f/f mouse exacerbates BRONJ‐like lesion. A) Fluorescence imaging of mitophagy events in flox or cKO BMDMs expressing mt‐Keima with or without ZA treatment for 6 h. The right panel shows quantitative measurements for the ratio of signal excited at 561 and 488 nm ( n = 20). Scale bars: 10 µm. B) Total protein expression of autophagic flux markers, and mitochondrial adaptor proteins in flox or cKO BMDMs with or without ZA treatment for 6 h. C,D) Panels show quantitative data of mean mtROS ( n = 3) and mean HIF‐1α expression ( n = 3) in flox or cKO BMDMs with or without ZA treatment. Scale bars: 5 µm. E) Total protein expression of HIF‐1α, iNOS, and CD206 in flox or cKO BMDMs with or without ZA treatment for 24 h. F–H) Panels show the quantitative data of open wound area ( n = 5), osteonecrosis area ( n = 5), trabecular bone volume/tissue volume (BV/TV) ( n = 5), and bone mineral density (BMD) ( n = 5). I) Flow cytometry assessment of jawbone macrophage stained with CD45, CD11b, F4/80, CD86, and CD206 antibodies. J) Immunofluorescence (IF) assay of F4/80 + CD86 + or F4/80 + CD206 + macrophages in TES. White arrows indicate representative cells. Scale bars: 100 µm. Results are representative of at least three independent experiments performed. Densitometric quantifications are indicated beneath the representative blots. Statistical significance was examined by a two‐sided one‐way ANOVA or Welch ANOVA (A,C,D,F–H). Results are presented as the mean ± SD.

Article Snippet: [ ] The cells were washed with PBS and stained with Fixable Viability Dye eFluor 506 (Invitrogen, 65‐0866‐14), APC‐eFlour 780 Anti‐Mouse CD45 (Invitrogen, 47‐0451‐82), PerCP/Cyanine5.5 Anti‐Mouse CD11b (Elabscience, E‐AB‐F1081J), Brilliant Violet 605 Anti‐Mouse F4/80 (Biolegend, 123133), PE Anti‐Mouse CD86 (BD Biosciences, 561963), and FITC Anti‐Mouse CD206 (Biolegend, 141703) antibodies.

Techniques: Fluorescence, Imaging, Expressing, Flow Cytometry, Staining, Immunofluorescence

Journal: Advanced Science

Article Title: Rescuing Mitochondrial Dysfunction in Macrophages Prevents Osteonecrosis of the Jaw in Anti‐Resorptive Therapy

doi: 10.1002/advs.202517586

Figure Lengend Snippet: Synthesis of ZDPR and its preventing effects of BRONJ. A) Schematic illustration of the preparation of ZDPR. B) Scanning electron microscopy (SEM) image showing the surface morphology of ZDPR nanoparticles. Scale bars: 100 nm. C) Drug‐releasing profile of RAPA in ZDPR nanoparticles. D–F) Western blot revealing total protein expression of mTOR, p‐mTOR, autophagic flux markers, mitochondrial adaptor proteins, HIF‐1α, pro IL‐1β, and IL‐1β in BMDMs treated with 5, 10, 15 µ m ZA or 5, 10, 15 µ m ZDPR for 6 or 24 h. G) Representative TEM images of BMDMs treated with ZA or ZDPR for 6 h. Red arrow indicates the typical formation of mitophagic vacuoles. White asterisks indicate damaged mitochondria. Zoomed boxed area presents a magnified structure of an early autophagic vacuole (AVi) containing a mitochondrion (mi). Scale bars: 500 nm. H) Fluorescence imaging of mitophagy events in BMDMs expressing mt‐Keima with ZA or ZDPR treatment for 6 h. Scale bars: 10 µm. I–K) Panels show the quantitative data of open wound area ( n = 5), osteonecrosis area ( n = 5), trabecular bone volume/tissue volume (BV/TV) ( n = 5), and bone mineral density (BMD) ( n = 5). L) Flow cytometry assessment of jawbone macrophage stained with CD45, CD11b, F4/80, CD86, and CD206 antibodies. M) Immunofluorescence (IF) assay of F4/80 + CD86 + or F4/80 + CD206 + macrophages in TES. White arrows indicate representative cells. Scale bars: 100 µm. Results are representative of at least three independent experiments performed. Densitometric quantifications are indicated beneath the representative blots. Panels show the quantitative data of mitophagy events per cell (G, n = 20), and the ratio of signal excited at 561 and 488 nm (H, n = 20). Statistical significance was examined by a two‐tailed Student's t ‐tests or two‐sided unpaired Welch's t ‐tests (G–K). Results are presented as the mean ± SD.

Article Snippet: [ ] The cells were washed with PBS and stained with Fixable Viability Dye eFluor 506 (Invitrogen, 65‐0866‐14), APC‐eFlour 780 Anti‐Mouse CD45 (Invitrogen, 47‐0451‐82), PerCP/Cyanine5.5 Anti‐Mouse CD11b (Elabscience, E‐AB‐F1081J), Brilliant Violet 605 Anti‐Mouse F4/80 (Biolegend, 123133), PE Anti‐Mouse CD86 (BD Biosciences, 561963), and FITC Anti‐Mouse CD206 (Biolegend, 141703) antibodies.

Techniques: Electron Microscopy, Western Blot, Expressing, Fluorescence, Imaging, Flow Cytometry, Staining, Immunofluorescence, Two Tailed Test

Journal: Biochemistry and Biophysics Reports

Article Title: HUC-MSC-derived exosomal miR-16-5p attenuates inflammation via dual suppression of M1 macrophage polarization and Th1 differentiation

doi: 10.1016/j.bbrep.2025.102078

Figure Lengend Snippet: miR-16-5p mimics blocks M1 macrophage polarization and Th1 cell differentiation. A: Raw264.7 cells were transfected with miR-16-5p mimics or NC mimics and subsequently induced to M1 macrophage polarization. After 24 h of culture, the percentage of CD11b + iNOS + cells in each group were measured by flow cytometry. B: Data are presented as mean ± standard deviation ( x ‾ ± s), with n = 3. p < 0.0001 compared with M1 group and NC mimics group. C: Naïve CD4 + T cells were induced to Th1 cells and then transfected with miR-16-5p or NC mimic. After 5 days of culture, the percentage of CD4 + IFN-γ + cells in each group were measured by flow cytometry. D: Data are presented as mean ± standard deviation ( x ‾ ±s), with n = 3. p = 0.0319, compared with Th1 group p = 0.0237, compared with NC mimics group. E: The inhibitory effect of miR-16-5p transfection on the mRNA levels of iNOS and TNF-α in M1 cells was measured by q-PCR. Data are presented as mean ± standard deviation ( x ‾ ±s), with n = 3. p iNOS <0.0001, p TNF-α = 0.0038, compared with M1 group, p iNOS <0.0001, p TNF-α = 0.0002, compared with NC mimics group. F: The mRNA expression levels of IFN-γ and STAT4 in different groups were measured by q-PCR. Data are presented as mean ± standard deviation ( x ‾ ±s), with n = 3. p STAT1 = 0.0018, p I - γ = 0.0032, compared with Th1 group, p STAT1 <0.0001, p I - γ = 0.0286, compared with NC mimics group.

Article Snippet: The single cells were labeled with iNOS (APC-iNOS, #17592082, eBioscience), F4/80 (FITC-F4/80, #35-4801-U100, Tonbo Biosciences), CD11b (PerCP-Cyanine5.5-CD11b, 65-0112-U100, Tonbo Biosciences), CD45(PE-CD45, #147712, Biolegend), CD3 (FITC-CD3, #35-0032-U100, Tonbo Biosciences) and CD4 (PE-CD4, #50-0041-U100, Tonbo Biosciences) respectively.

Techniques: Cell Differentiation, Transfection, Flow Cytometry, Standard Deviation, Expressing

Journal: Biochemistry and Biophysics Reports

Article Title: HUC-MSC-derived exosomal miR-16-5p attenuates inflammation via dual suppression of M1 macrophage polarization and Th1 differentiation

doi: 10.1016/j.bbrep.2025.102078

Figure Lengend Snippet: miR-34a-5p and miR-125b-5p mimics suppress M1 macrophage polarization and Th1 cell differentiation. A: The Raw264.7 cells were transfected miR-34a-5p mimic, miR-125b-5p mimics or NC mimics and subsequently induced to M1 macrophage. After 24 h of culture, the percentage of CD11b + iNOS + cells in various group was measured by flow cytometry. B: Data are presented as mean ± standard deviation ( x ‾ ± s), with n = 3. p M1,34 = 0.0118, p M1,125 = 0.0037, compared with M1 group; p NC,34 = 0.0202, p NC,125 = 0.0060, compared with NC mimics group. C: Naïve CD4 + T cells were induced to Th1 cells and then transfected with miR-34a-5p mimics, miR-125b-5p mimics or NC mimic. After 5 days of culture, the percentage of CD4 + IFN-γ + cells in various groups was measured by flow cytometry. D: Data are presented as mean ± standard deviation ( x ‾ ± s), with n = 3. p Th1,34 = 0.0102, p Th1,125 = 0.0201, compared with Th1 group, p NC,34 = 0.0220, p NC,125 = 0.0447, compared with NC mimics group. E: The inhibitory effects of miR-34a-5p mimics or miR-125b-5p mimics transfection on the mRNA levels of iNOS and TNF-α in M1 cells was measured by q-PCR. Data are presented as mean ± standard deviation ( x ‾ ± s), with n = 3. p 34-iNOS <0.0001, p 125-iNOS <0.0001, p 34-TNF-α = 0.0012, p 125-TNF-α = 0.0117, compared with M1 group, p 34-iNOS = 0.0001, p 125-iNOS = 0.0027, p 34-TNF-α = 0.0039, p 125-TNF-α = 0.0377, compared with NC mimics group. F: The mRNA expression level of I - γ and STAT1 in different groups were measured by q-PCR. x ‾ ± s, with n = 3. p 34-STAT1 <0.0001, p 125-STAT1 <0.0001, p 3 - γ <0.0001, p 1 - γ <0.0001, compared with Th1 group, p 34-STAT1 <0.0001, p 125-STAT1 <0.0001, p 3 - γ = 0.0014, p 1 - γ = 0.0004, compared with NC mimics group.

Article Snippet: The single cells were labeled with iNOS (APC-iNOS, #17592082, eBioscience), F4/80 (FITC-F4/80, #35-4801-U100, Tonbo Biosciences), CD11b (PerCP-Cyanine5.5-CD11b, 65-0112-U100, Tonbo Biosciences), CD45(PE-CD45, #147712, Biolegend), CD3 (FITC-CD3, #35-0032-U100, Tonbo Biosciences) and CD4 (PE-CD4, #50-0041-U100, Tonbo Biosciences) respectively.

Techniques: Cell Differentiation, Transfection, Flow Cytometry, Standard Deviation, Expressing