counterstaining with dapi  (Beyotime)

Journal: bioRxiv

Article Title: Age-related loss of brain demyelinating and remyelinating potential is overcome by microglia renewal

doi: 10.64898/2026.06.15.732390

Figure Lengend Snippet: Age-dependent impairment in myelin phagocytosis and lipid droplet clearance in vitro and in vivo (Ai-ii) In vitro myelin phagocytosis assay with primary peritoneal macrophages from young (1-month-old) and aged (12-month-old) mice. (Ai) Representative immunofluorescence cytochemistry images (myelin basic protein (MBP, green), Iba1 (red), and DAPI (blue) of macrophages from young and aged mice incubated with myelin debris for 2, 4, 16, or 24 h. Scale bar: 20μm. (Aii) Quantification of macrophages containing phagocytosed myelin (% of MBP-positive Iba1- macrophages) at early (2–4 h) and late (16–24 h) time points. (Bi-iii) Delayed metabolism of phagocytosed myelin by aged macrophages. (Bi) Experimental scheme showing removal of myelin from culture medium after 16 h, and assessment of intracellular myelin metabolism by accumulation of Oil-Red-O (ORO) lipid droplets at 24 and 48 h. As an experimental control, representative images of MBP (green), Iba1 (red), and DAPI (blue) (Scale bar: 20μm) in young and aged macrophages showing comparable levels of phagocytosed MBP-immunoreactive myelin in young and aged macrophages at 16 h, and absence of MBP-immunoreactive myelin at 24 h and 48 h, (Bii) ORO and hematoxylin staining of young and aged macrophages showing ORO+ lipid droplet accumulation (red, arrowheads). (Biii) Quantification of ORO+ lipid–containing macrophages following myelin incubation and withdrawal, demonstrating impaired lipid clearance in aged compared to young macrophages. Scale bar: 3μm. (Ci-iii) In vivo accumulation of ORO+ lipid droplets in young (2-month-old) and old (24-month-old) mice during cuprizone-induced demyelination. (Ci) Representative ORO-stained coronal brain sections from young and aged mice after 4 (CPZ4), 6 (CPZ6) and 8 (CPZ8) weeks of continuous cuprizone feeding, and CPZ6 followed by 4 weeks of recovery after cuprizone withdrawal (CPZ6+4). Scale bar: 10μm. Quantification of ORO+ lipid droplets in, (Cii) the corpus callosum (CC) and (Ciii) dorsal fornix (DF) between young and aged mice at CPZ0, CPZ4, CPZ6, CPZ6+4 and CPZ8. (n = 5 mice/group; Mean ± SEM, p<0.05, *p<0.01, **p<0.001, ***p<0.0001).

Article Snippet: This was followed by a three-probe amplification steps (AMP1-AMP3), fluorophore conjugation (Opal 570) and DAPI counterstaining (ACD, CA).

Techniques: In Vitro, In Vivo, Phagocytosis Assay, Immunofluorescence, Incubation, Control, Staining

Journal: Bioengineering & Translational Medicine

Article Title: Extracellular matrix microarchitecture modulates cellular behavior and extracellular vesicle phenotypes in biomimetic tendon models

doi: 10.1002/btm2.70134

Figure Lengend Snippet: (a) Actin (green) and DAPI (blue) stain of tendon derived cells seeded on tissue culture plate (TCP) monolayer controls, healthy mimetic models and diseased mimetic models ( n = 3). Scale bar: 100 μm. (b) Nuclear aspect ratio (NAR) was quantified using ImageJ by converting microscopic image to grayscale and measuring major and minor axes of individual nuclei. A higher major‐to‐minor ratio indicates increased cellular elongations. (c) Cells on the healthy model showed greater elongation than both the monolayer control and diseased model by day 3, while the diseased model also exceeded the monolayer control. By day 5, elongation remained elevated only in the healthy model relative to the diseased, and by day 7, the healthy group exhibited significantly greater elongation than all other models. No significant differences were observed by day 10. Significance was defined as p < 0.05 (*), p < 0.01 (**), p < 0.001 (***), p < 0.0001 (****), ns = not significant.

Article Snippet: Cellular morphology was assessed via actin staining (Invitrogen, Cat. No. A12379) and nuclear counterstaining with DAPI (Fisher Scientific, Cat. No. H‐1200‐10) ( n = 3).

Techniques: Staining, Derivative Assay, Control