mab1556  (R&D Systems)

Journal: Nature Physics

Article Title: Boundary-guided cell alignment drives mouse epiblast maturation

doi: 10.1038/s41567-026-03176-9

Figure Lengend Snippet: ( A ) Box plots of EPI cell number, showing median (centre), 25th–75th percentiles (box), with whiskers extending to 1.5x interquartile range, with individual embryos shown as dots. Sample sizes: Lamc1 +/+ , n = 9 (E4.5), 13 (E4.75), 11 (E5.0); Lamc1 +/− , n = 12, 28, 20; Lamc1 −/− , n = 4, 11, 9 embryos from 5, 7, and 5 independent Lamc1 +/− × Lamc1 +/− litters. One-way ANOVA with Tukey’s post hoc test, ns = not significant. ( B ) Box plots of EPI cell number, as in ( A ). Sample sizes: Itgb1 +/+ , n = 5 (E4.5), 13 (E4.75), 9 (E5.0); Itgb1 +/− , n = 13, 28, 26; Itgb1 −/− , n = 8, 14, 11 embryos from 5, 8, and 7 independent Itgb1 +/− × Itgb1 +/− litters. One-way ANOVA with Tukey’s post hoc test. *p < 0.05, **p < 0.01, ns = not significant. At E5.0, Itgb1 +/+ vs Itgb1 −/− , p = 0.017; Itgb1 +/− vs Itgb1 −/− , p = 0.001. ( C ) Immunofluorescence images of representative Lamc1 +/+ or +/− and Lamc1 −/− embryos at E5.25, stained for Oct3/4 (EPI, cyan), phalloidin (yellow), and podocalyxin (magenta). n = 28 ( Lamc1 +/+ or +/− ), 6 ( Lamc1 −/− ) embryos from 4 independent Lamc1 +/− × Lamc1 +/− litters. ( D ) Immunofluorescence images of representative Itgb1 +/+ or +/− and Itgb1 −/− embryos at E5.25, stained for Oct3/4 (EPI, cyan), pan-laminin (green), and phalloidin (yellow). n = 13 ( Itgb1 +/+ or +/− ), 2 ( Itgb1 −/− ) embryos from 2 independent Itgb1 +/− × Itgb1 +/− litters. ( E ) Long axis length measurement, shown as violin plots with individual data points. Dot colours indicate embryo stage; red bars show medians. Sample sizes: Lamc1 +/+ , n = 163 cells from 6 embryos (E4.5), 397 from 8 (E4.75), 571 from 10 (E5.0); Lamc1 +/− , n = 199 from 7, 583 from 14, 637 from 10; Lamc1 − / − , n = 75 from 3, 277 from 7, 462 from 9. Mann-Whitney U test (two-sided) without correction for multiple comparisons, each group compared to the reference group [15-29 cells]. *p < 0.05, **p < 0.01, ***p < 0.001, ns = not significant. Exact p-values: Lamc1 +/+ [30–44], p = 0.345; [45–59], p = 0.189; [60–74], p = 0.236; [75–89], p = 8.72 × 10 −9 . Lamc1 +/− [30–44], p = 0.051; [45–59], p = 1.00 × 10 −3 ; [60–74], p = 5.02 × 10 −6 ; [75–89], p = 7.42 × 10 −6 . Lamc1 −/− [30–44], p = 0.539; [45–59], p = 4.07 × 10 −4 ; [60–74], p = 0.059. ( F ) Long axis length measurement, as in ( E ). Sample sizes: Itgb1 +/+ , n = 110 cells from 3 embryos (E4.5), 399 from 8 (E4.75), 251 from 4 (E5.0); Itgb1 +/− , n = 200 from 5, 560 from 11, 618 from 10; Itgb1 −/− , n = 209 from 7, 470 from 11, 360 from 9. Mann-Whitney U test (two-sided) without correction for multiple comparisons, each group compared to the reference group [15-29 cells]. *p < 0.05, **p < 0.01, ***p < 0.001, ns = not significant. Exact p-values: Itgb1 +/+ : [30–44], p = 2.36 × 10 −4 ; [45–59], p = 0.015; [60–74], p = 8.97 × 10 −3 ; [75–89], p = 2.34 × 10 −4 . Itgb1 +/− : [30–44], p = 0.845; [45–59], p = 0.335; [60–74], p = 0.518; [75–89], p = 0.022. Itgb1 −/− : [30–44], p = 5.65 × 10 −4 ; [45–59], p = 0.022; [60–74], p = 3.71 × 10 −4 . Scale bars, 20 µm. See also Fig. .

Article Snippet: Primary antibodies against laminin (Novus-Biologicals, NB300-144), pERM (Cell Signaling, 3726) and podocalyxin (R&D Systems, MAB1556) were diluted at 1:200.

Techniques: Immunofluorescence, Staining, MANN-WHITNEY

Journal: Glia

Article Title: LRP4+ Astrocytes: A Unique Subpopulation Crucial for Blood Vessel Maintenance and Function in the Somatosensory Cortex of Normal and 5xFAD Mice

doi: 10.1002/glia.70114

Figure Lengend Snippet: Lrp4+ astrocytes make direct contact with the pia and cortical surface arteries and exhibit robust soma‐artery interactions. (A) Tissue cutting schematic. Lrp4creER; Ai9 mice were used for whole‐mount transverse tissue sectioning for cortical surface immunostaining. Mice were administered 100 mg/kg TAM at 2/3 MO and sacrificed a month later. Tissue sections were ~1 mm deep into the cortex and contained layers of the meninges such as the pia. (B) Representative 3D reconstructions of the cortical surface based on IHC analysis of the pia (Laminin‐2), tdTom+, and GFAP+ cells. Reconstructions are paired with respective single plane confocal images of each layer. Scale bar = 60 μm. (C) Representative 3D reconstructions of the cortical surface based on IHC analysis of SMA‐positive artery, tdTom+, and GFAP+ cells at the cortical surface. (D) IHC analysis of smooth muscle actin (SMA)‐positive arteries with tdTom+ and GFAP+ cells. Representative images from the cortical surface are shown. Scale bar = 100 μm. (E) IHC analysis of podocalyxin (Podxl)‐positive blood vessels with tdTom+ and GFAP+ cells. Representative images from the cortical surface are shown. Scale bar = 45 μm. (F) Quantification of data in panels (D and E), respectively. Percent (%) of total SMA‐positive artery (D) and Podxl‐positive blood vessel (E) area covered by GFAP+ and tdTom+ cells. (G) Quantification of data in panel (D). Quantification of percentage (%) of total tdTom+ and GFAP+ cells interacting with a SMA‐positive artery by either the whole cell body, or by processes only. n = 5–7. Paired t tests. * p < 0.05, **** p < 0.0001.

Article Snippet: The following primary antibodies were used for immunostaining: IBA1 (abcam Cat# ab5076), OLIG2 (invitrogen Cat# p21954), GFAP (Thermofisher Catalog # PA1‐10004), CRABP2 (abcam Cat#: ab211927), Laminin‐2 (Sigma #L0663), SMA (santa cruz biotechnology Cat#: sc‐3225), Podxl (R&D Systems Ca# MAB1556), PDGFRβ (abcam Cat#: ab 32570), CD11b (abcam Cat#: ab8878), 6E10 (BioLegend Ca# 803020), Laminin‐α5 (Abcam Cat#: ab17107).

Techniques: Immunostaining

Journal: Glia

Article Title: LRP4+ Astrocytes: A Unique Subpopulation Crucial for Blood Vessel Maintenance and Function in the Somatosensory Cortex of Normal and 5xFAD Mice

doi: 10.1002/glia.70114

Figure Lengend Snippet: Loss of Lrp4+ astrocytes results in decreased blood vessel density, branching, diameter and blood flow at the cortical surface. (A) IHC analysis of Podxl+ blood vessels (BVs) in coronally sectioned tissue from iDTR control and ablation mice. Representative image of the pia is shown here. Scale bar = 30 μm. (B) Quantification of data in panel (A) comparing density per mm 2 and (C) diameter (μm) of BVs in the pia of iDTR Control and Ablation mice. (D) IHC analysis of SMA+ arteries and Podxl+ blood vessels (BVs) in transverse sectioned tissue from iDTR control and ablation mice. Representative image of the cortical surface are shown here. Scale bar = 250 μm. (E) Quantification of data in panel (D) comparing artery diameter (μm), (F) BV density (mm 2 ), and (G) # of BV branches (mm 2 ) in iDTR control and ablation mice. (H) Representative image of unilateral cerebral blood flow analyses via laser speckle contrast imaging. (I) Quantification of average recorded perfusion (PU) of iDTR Control and Ablation mice. n = 3–4. Unpaired t ‐tests. Data are mean ± SEM. ** p < 0.01, *** p < 0.001, **** p < 0.0001.

Article Snippet: The following primary antibodies were used for immunostaining: IBA1 (abcam Cat# ab5076), OLIG2 (invitrogen Cat# p21954), GFAP (Thermofisher Catalog # PA1‐10004), CRABP2 (abcam Cat#: ab211927), Laminin‐2 (Sigma #L0663), SMA (santa cruz biotechnology Cat#: sc‐3225), Podxl (R&D Systems Ca# MAB1556), PDGFRβ (abcam Cat#: ab 32570), CD11b (abcam Cat#: ab8878), 6E10 (BioLegend Ca# 803020), Laminin‐α5 (Abcam Cat#: ab17107).

Techniques: Control, Imaging

Journal: Glia

Article Title: LRP4+ Astrocytes: A Unique Subpopulation Crucial for Blood Vessel Maintenance and Function in the Somatosensory Cortex of Normal and 5xFAD Mice

doi: 10.1002/glia.70114

Figure Lengend Snippet: Loss of Lrp4+ astrocytes results in decreased blood vessel density, branching and diameter, along with decreased pericyte and laminin‐α5 coverage. (A) IHC analysis of Podxl+ blood vessels (BVs) in coronally sectioned tissue from iDTR control and ablation mice. Representative images contain cortical layers 1–4. Area indicated by white boxes are presented at higher resolution in bottom panels. Scale bar = 50 μm. (B) Quantification of BV diameter (μm), (C) density (mm 2 ), and (D) # of BV branches (per mm 2 ) in iDTR Control and Ablation mice in L1‐4 of the ssc. Unpaired t ‐tests. * p < 0.05, ** p < 0.01, **** p < 0.0001. (E) IHC analysis of pericytes (Pdgfrβ) and Podxl+ BVs from the same tissue used in panel (A). Representative image is from cortical layer 3. White arrows indicate BV‐associated pericytes. Scale bar = 35 μm. (F) Quantification of pericyte density (# pericytes per mm 2 ) in iDTR control and ablation mice. n = 4–5. Data are mean ± SEM. Unpaired t ‐test. * p < 0.05, ** p < 0.01, **** p < 0.0001. (G) IHC of Podxl+ BVs and Caspase‐3 along the pia and in L1‐4 cortex of iDTR control and ablation mice. Pia scale bar = 50 μm. Cortex scale bar = 35 μm. (H) Quantification of percent (%) BV covered by Caspase‐3 (Caspase‐3 + Podxl/total Podxl) in iDTR control and ablation mice. n = 4–5. (I) IHC of Podxl+ BVs and Collagen V along the pia of iDTR control and ablation mice. Scale bar = 50 μm. (J) Quantification of percent (%) BV covered by Collagen V (Collagen V + Podxl/total Podxl) in iDTR control and ablation mice. n = 3. (K) IHC of Podxl+ BVs and Laminin‐α5 at the pia and cortex L3 of iDTR control and ablation mice. Scale bar = 20 μm. (L) Quantification of percent (%) BV covered by Laminin‐α5 (Laminin‐α5 + Podxl/total Podxl) at both the pia and cortex of iDTR control and ablation mice. n = 3. Data are mean ± SEM. Unpaired t ‐test. **** p < 0.0001. (M) IHC of Laminin‐α5 with GFAP, IBA1, and LRP4+ astrocytes (tdTom) in cortex L3 of iDTR control and ablation mice. Scale bar = 25 μm. (N) Quantification of percent (%) area of GFAP+, IBA1+, and LRP4+ cells colocalized with the Laminin‐α5. n = 3. Data are mean ± SEM. Multiple t ‐tests. **** p < 0.0001.

Article Snippet: The following primary antibodies were used for immunostaining: IBA1 (abcam Cat# ab5076), OLIG2 (invitrogen Cat# p21954), GFAP (Thermofisher Catalog # PA1‐10004), CRABP2 (abcam Cat#: ab211927), Laminin‐2 (Sigma #L0663), SMA (santa cruz biotechnology Cat#: sc‐3225), Podxl (R&D Systems Ca# MAB1556), PDGFRβ (abcam Cat#: ab 32570), CD11b (abcam Cat#: ab8878), 6E10 (BioLegend Ca# 803020), Laminin‐α5 (Abcam Cat#: ab17107).

Techniques: Control