Journal: Exploration

Article Title: RBFOX1 Dysfunction Unlocks APOE4 ‐Associated Microglial Genesis and Exacerbates Alzheimer's Pathology in Human Cerebral Organoids

doi: 10.1002/exp2.70160

Figure Lengend Snippet: RBFOX1 knockout triggers APOE4 ‐specific microglia generation in human cerebral organoids. (A) Schematic of generation of organoids, (B) Generation of APOE3 , APOE3_RBFOX1 ‐KO, APOE4 , and APOE4_RBFOX1 ‐KO H9 cell lines using CRISPR/Cas9 technology, (C) Representative bright‐field images (Day 60, Scale bars, 500 µm), (D) Immunostaining for PAX6 (red), TUJ1 (green), and DNA (Hoechst, blue) (Day 60, Scale bars, 50 µm) showed organoids well developed. (E) Gene expression heatmap of microglia signature genes in APOE3 , APOE3_RBFOX1 ‐KO, APOE4 , and APOE4_RBFOX1 ‐KO organoids at Day 60 from RNA‐seq data. (two biological replicates per genotype, with three organoids pooled per replicate). A pseudo‐count was used for FPKM values (FPKM + 1), log2‐transformed, and each gene was normalized in its respective row. (F, G) Representative images showing the microglia marker IBA1 and TMEM119 in organoids at Day 60, respectively. Scale bars, 50 µm, (H, I) Quantification of IBA1 and TMEM119 average fluorescence intensity shown in (F, G) respectively. Data represent the mean ± SEM and were analyzed by two‐way ANOVA. ( n = 10; each data point represents one organoid; organoids were collected from three independent batches; image fields were randomly selected, with a minimum of eight fields analyzed per replicate), and (J) Flow cytometry analysis of IBA1 in Day 60 organoids.

Article Snippet: Primary antibodies used in this study included rabbit IBA1 (1:400, Wako, 019–19741), rabbit TMEM119 (1:400, Abcam, ab185333), rat CD68 (1:200, Invitrogen, 14‐0681‐82), mouse PSD95 (1:400, Abcam, ab2723), rabbit synapsin1 (1:400, Abcam, ab64581), mouse Tuj1 (1:400, Abcam, ab78078), mouse Pax6 (1:200, DSHB, AB_528427), mouse PHF1 (1:1000, Santa Cruz Biotechnology, SC‐81376), mouse AT8 (1:400, Pierce, mn1020), mouse 6E10 (1:400, Covence, SIG‐39300), and mouse 4G8 (1:400, Covence, SIG‐39200), LipidTOX (1:1000, Invitrogen, H34477 ).

Techniques: Knock-Out, CRISPR, Immunostaining, Gene Expression, RNA Sequencing, Transformation Assay, Marker, Fluorescence, Flow Cytometry

Journal: Frontiers in Molecular Neuroscience

Article Title: Cohesive Regulation of Neural Progenitor Development by microRNA miR-26, Its Host Gene Ctdsp and Target Gene Emx2 in the Mouse Embryonic Cerebral Cortex

doi: 10.3389/fnmol.2018.00044

Figure Lengend Snippet: Ctdsp2 increases neural progenitor (NP) proliferation. (A–C) Overexpression of Ctdsp2 , but not the control construct pCAGIG, increased the proportion of cells expressing both proliferative marker bromodeoxyuridine + (BrdU + )/green fluorescence protein + (GFP + ) and radial glial cell (RGC) marker Pax6 + /GFP + , but not intermediate progenitor (IP) marker Tbr2 + /GFP + , in GFP-positive cells. (D–F) shRNA-mediated knockdown (sh Ctdsp2 ) of Ctdsp2 decreased the proportion of both BrdU + /GFP + cells and Pax6 + /GFP + cells, but not Tbr2 + /GFP + cells in GFP-positive cells, compared to the control construct pSilencer. Values represent mean ± SEM. n = 9 sections from at least three brains. ** P < 0.01; ns, not significant. ANOVA with post hoc test was used. Scale bar = 50 μm.

Article Snippet: Primary antibodies against the following antigens were used: BrdU (1:50, Developmental Studies Hybridoma Bank at University of Iowa (DSHB)), Ki67 (1:500, Abcam), Pax6 (1:500, Covance), Pax6 (1:15 DSHB), Tbr2 (1:500, Abcam), GFP (1:1000, Abcam, chicken), and GFP (1:1000, Rockland, rabbit).

Techniques: Over Expression, Control, Construct, Expressing, Marker, Fluorescence, shRNA, Knockdown

Journal: Frontiers in Molecular Neuroscience

Article Title: Cohesive Regulation of Neural Progenitor Development by microRNA miR-26, Its Host Gene Ctdsp and Target Gene Emx2 in the Mouse Embryonic Cerebral Cortex

doi: 10.3389/fnmol.2018.00044

Figure Lengend Snippet: miR-26 promotes NP proliferation. (A,D,G) Overexpression of miR-26a, but not the control construct pCAGIG, proportion of cells expressing both proliferative marker BrdU + /GFP + and RGC marker Pax6 + /GFP + , but not IP marker Tbr2 + /GFP + , in GFP-positive cells. (B,E,H) miRNA sponge-mediated knockdown (miR-26-SP), but not the mutated sponge (miR-26-SPmut), decreased proportion of cells expressing both proliferative marker BrdU + /GFP + and RGC marker Pax6 + /GFP + in GFP-positive cells. (C,F,I) Ratio of BrdU + /GFP + , Pax6 + /GFP + or Tbr2 + /GFP + cells vs. GFP + cells in the electroporated cortex. Values represent mean ± SEM. n = 9 sections from at least three brains. * P < 0.05; ** P < 0.01; ns, not significant. ANOVA with post hoc test was used. Scale bar = 50 μm.

Article Snippet: Primary antibodies against the following antigens were used: BrdU (1:50, Developmental Studies Hybridoma Bank at University of Iowa (DSHB)), Ki67 (1:500, Abcam), Pax6 (1:500, Covance), Pax6 (1:15 DSHB), Tbr2 (1:500, Abcam), GFP (1:1000, Abcam, chicken), and GFP (1:1000, Rockland, rabbit).

Techniques: Over Expression, Control, Construct, Expressing, Marker, Knockdown

Journal: Frontiers in Molecular Neuroscience

Article Title: Cohesive Regulation of Neural Progenitor Development by microRNA miR-26, Its Host Gene Ctdsp and Target Gene Emx2 in the Mouse Embryonic Cerebral Cortex

doi: 10.3389/fnmol.2018.00044

Figure Lengend Snippet: Emx2 is functionally inhibited by miR-26 in regulating NP proliferation. (A–F) Overexpression of Emx2 , but not the control construct pCAGIG, decreased the proportion of cells expressing both proliferative marker BrdU + /GFP + and RGC marker Pax6 + /GFP + , but not IP marker Tbr2 + /GFP + , in GFP-positive cells. short hairpin RNA (shRNA)-mediated knockdown (sh Emx2 ) of Emx2 increased the proportion of both BrdU + /GFP + cells and Pax6 + /GFP + cells, but not Tbr2 + /GFP + cells in GFP-positive cells, compared to the control construct pSilencer. (G–J) Emx2 expression suppressed the proportion of cells expressing both proliferative marker BrdU + /GFP + and RGC marker Pax6 + /GFP + in GFP-positive cells. Co-expressing Emx2 with miR-26, but not miR-26-mut, dramatically reversed the suppression. (K,L) Emx2 expression did not alter the proportion of cells expressing IP marker Tbr2 + /GFP + , in GFP-positive cells. Values represent mean ± SEM. n = 9 sections from at least three brains. ** P < 0.01; *** P < 0.001; ns, not significant. ANOVA with post hoc test was used. Scale bar = 50 μm.

Article Snippet: Primary antibodies against the following antigens were used: BrdU (1:50, Developmental Studies Hybridoma Bank at University of Iowa (DSHB)), Ki67 (1:500, Abcam), Pax6 (1:500, Covance), Pax6 (1:15 DSHB), Tbr2 (1:500, Abcam), GFP (1:1000, Abcam, chicken), and GFP (1:1000, Rockland, rabbit).

Techniques: Over Expression, Control, Construct, Expressing, Marker, shRNA, Knockdown

Journal: Cell stem cell

Article Title: PTP1B is an effector of activin signaling and regulates neural specification of embryonic stem cells.

doi: 10.1016/j.stem.2013.09.016

Figure Lengend Snippet: Figure 7. PTP1B Promotes Mesendodermal Fates and Restricts Neural Differentiation (A) PTP1B hESC mutants were verified by western blot and their effects on the levels of p-ERK1/2. (B) The expression profiles of fate markers in PTP1B mutant hESCs as assessed by RT-PCR. (C) Three PTP1B-KD hESC clones were subjected to neural differentiation in the absence of SB431542, and PAX6+ cells (red) were counted. Data are presented as mean ± SD (n = 3). **p < 0.01, significantly different from the control group. (D) PTP1B-eGFP-overexpressing hESCs favor mesendodermal fate, judging by SOX17 expression. (E) A schematic diagram depicts the conceptual summary derived from this study. Scale bars represent 50 mm. See also Figure S7.

Article Snippet: Antibodies against Oct4 (1:500, AB3209, Chemicon), Nes (1:500, MAB353, Chemicon), Pax6 (1:500, Hybridoma bank), PTP1B (1:500, Santa Cruz Biotechnology), Alk4 (1:500, Santa Cruz Biotechnology), and NeuN (1:500, MAB377, Chemicon) were used.

Techniques: Western Blot, Expressing, Mutagenesis, Reverse Transcription Polymerase Chain Reaction, Clone Assay, Control, Derivative Assay

Journal: Nature Neuroscience

Article Title: Single-cell genotyping and transcriptomic profiling of mosaic focal cortical dysplasia

doi: 10.1038/s41593-025-01936-z

Figure Lengend Snippet: a , UMAP visualization comparing age distribution of subjects between the reference Velmeshev et al. postmortem control dataset (V19, left) and the complete integrated dataset from this study (right). Color gradient indicates subject age in years. b , Relative proportion of major cell types across individual samples. Cell types include glutamatergic neurons (GluN and GluL2-6), GABAergic interneurons (IN-MGE and IN-CGE), glial cells (astrocytes, oligodendrocytes, OPCs) and other cell types (microglia, endothelial cells). Abbreviations: Glu, glutamatergic; N, neurons; L, layer; CC, cortico-cortical projection neurons; IN-MGE/CGE, interneurons originating from the medial/caudal ganglionic eminence; OPC, oligodendrocyte precursor cells. c , Individual UMAP plots showing nucleus distribution for each patient and control. d , Quality metrics for snRNA-seq data across cell types and individuals: total count of unique molecular identifiers (UMIs) per nucleus (N counts), mean number of unique genes (N genes) detected per nucleus and percentage (%) of transcripts from mitochondrial genes.

Article Snippet: Nuclei were resuspended in staining buffer (2% bovine serum albumin, 1 mM EDTA and phosphate-buffered saline) and immunostained overnight at 4 °C with the following primary antibodies: conjugated anti-NEUN-PE for neurons (1:1,000, Milli-Mark, cat. no. FCMAB317PE) , conjugated anti-PU.1-AF647 for microglia (1:100, Cell Signaling Technology, cat. no. 2240S conjugate) , , unconjugated anti-OLIG2 for oligodendrocytes (1:500, Abcam, cat. no. ab109186) , conjugated anti-PAX6-APC for astrocytes (1:1,000, Novus Biologicals, cat. no. NBP2-34705APC) and rabbit unconjugated anti-TBR1 for excitatory neurons (1:1,000, Abcam, cat. no. ab31940) .

Techniques: Control

Journal: Nature Neuroscience

Article Title: Single-cell genotyping and transcriptomic profiling of mosaic focal cortical dysplasia

doi: 10.1038/s41593-025-01936-z

Figure Lengend Snippet: a , Quantitative assessment of differentially expressed genes (DEGs) between patients and controls (log 2 (FC) > 0.4, p-value < 0.05, Fig. ). Left: Correlation between DEG count and total number of nuclei per cell type. Right: Correlation between DEG count and mean number of genes detected per nucleus in each cell type. Pearson correlation coefficients (R) and p-values are shown. Abbreviations: Astro, astrocytes; Glu, glutamatergic; N, neurons; L, layer; IN-MGE/CGE, interneurons originating from the medial/caudal ganglionic eminence; Micro, microglia; Oligo, oligodendrocytes; OPC, oligodendrocyte precursor cells. b-f , Gene Ontology (GO) pathway analysis of DEGs comparing focal FCDII patients to controls for interneurons ( b ), glutamatergic neurons ( c ), astrocytes ( d ), oligodendrocytes ( e ) and microglia ( f ). Top 10 significantly enriched GO terms with adjusted (adj.) p-value < 0.05 are shown per cell type.

Article Snippet: Nuclei were resuspended in staining buffer (2% bovine serum albumin, 1 mM EDTA and phosphate-buffered saline) and immunostained overnight at 4 °C with the following primary antibodies: conjugated anti-NEUN-PE for neurons (1:1,000, Milli-Mark, cat. no. FCMAB317PE) , conjugated anti-PU.1-AF647 for microglia (1:100, Cell Signaling Technology, cat. no. 2240S conjugate) , , unconjugated anti-OLIG2 for oligodendrocytes (1:500, Abcam, cat. no. ab109186) , conjugated anti-PAX6-APC for astrocytes (1:1,000, Novus Biologicals, cat. no. NBP2-34705APC) and rabbit unconjugated anti-TBR1 for excitatory neurons (1:1,000, Abcam, cat. no. ab31940) .

Techniques:

Journal: Nature Neuroscience

Article Title: Single-cell genotyping and transcriptomic profiling of mosaic focal cortical dysplasia

doi: 10.1038/s41593-025-01936-z

Figure Lengend Snippet: a , Distribution of 808 genotyped nuclei in UMAP space: 117 were classified as Mut. (pt10 = 89, pt9 = 25, pt7 = 2, pt6 = 1) or as Ref. Right, Mut. nuclei percentages per cell type (top) and across cell types (bottom). b , Representative images of co-immunofluorescence staining on formalin-fixed paraffin-embedded sections ( n = 1 per patient) showing mTOR-hyperactive (pS6 + ) neurons (NEUN + , pt2), astrocytes (GFAP + , pt2), oligodendrocytes (OLIG2 + , pt2) and microglia (IBA1 + , pt10). Nuclei (in blue) are labeled with DAPI. Scale bars, 20 µm. All patients included in this experiment are detailed in Supplementary Table . c , Cytomegalic cells representing a minor fraction of mutated cells. Left, representative immunostaining of SMI311 + DNs and VIM + BCs on frozen brain tissue from pt5. Nuclei (in blue) are labeled with DAPI for total cell counting. Scale bar, 25 µm. Right, mutated cell percentage (inferred by the detected VAF) and proportion of DNs or BCs identified in each patient ( n = 1 section/patient/staining was analyzed). d , Schematic of the distribution of mutated cells across cell types and the fraction of mutated cytomegalic cells in pt10. Astro, astrocytes; Endo, endothelial cells; Hemi, hemispherical; Oligo, oligodendrocytes; Micro, microglia.

Article Snippet: Nuclei were resuspended in staining buffer (2% bovine serum albumin, 1 mM EDTA and phosphate-buffered saline) and immunostained overnight at 4 °C with the following primary antibodies: conjugated anti-NEUN-PE for neurons (1:1,000, Milli-Mark, cat. no. FCMAB317PE) , conjugated anti-PU.1-AF647 for microglia (1:100, Cell Signaling Technology, cat. no. 2240S conjugate) , , unconjugated anti-OLIG2 for oligodendrocytes (1:500, Abcam, cat. no. ab109186) , conjugated anti-PAX6-APC for astrocytes (1:1,000, Novus Biologicals, cat. no. NBP2-34705APC) and rabbit unconjugated anti-TBR1 for excitatory neurons (1:1,000, Abcam, cat. no. ab31940) .

Techniques: Immunofluorescence, Staining, Formalin-fixed Paraffin-Embedded, Labeling, Immunostaining, Cell Counting

Journal: Nature Neuroscience

Article Title: Single-cell genotyping and transcriptomic profiling of mosaic focal cortical dysplasia

doi: 10.1038/s41593-025-01936-z

Figure Lengend Snippet: a , Expression analysis of cell-type-specific markers in mutation detected (Mut.) and reference detected (Ref.) nuclei. Dot size represents the proportion of nuclei expressing each marker; color intensity indicates average normalized expression level. Abbreviations: N, neurons; Glut.N, glutamatergic neurons; IN, interneurons; Astro, astrocytes; Oligo, oligodendrocytes; Micro, microglia. b , Fluorescence-activated nuclei sorting (FANS) gating strategy for cell population enrichment. Representative gating from pt9 is shown. Sequential gating begins with initial selection based on DAPI nuclear staining, followed by separation of neuronal (NEUN+) and non-neuronal populations. Cell-type-specific enrichment was then achieved using TBR1 for glutamatergic neurons, PAX6+/NEUN- for astrocytes, OLIG2+/NEUN- for oligodendrocytes, and PU.1 + /NEUN- for microglia. Note: TBR1 subpopulation analysis was only performed for pt9 due to tissue constraints. c , Quantification of somatic mutations across FANS-enriched cell populations was performed using two complementary approaches: ddPCR detection for MTOR and PIK3CA variants, and deep targeted amplicon sequencing (TAS) for the RHEB variant in pt9. Mutation-positive bulk brain DNA and mutation-negative blood DNA served as controls. ddPCR detection limits (LOD) were >3 FAM+ mutated droplets for MTOR , >6 FAM+ mutated droplets for PIK3CA in ddPCR analysis. Results are presented as mean ± SD where technical replication was feasible. Due to limited tissue availability, biological replicates could not be performed, and technical replicates were not possible for specific cell populations in patients pt4 (PAX6+/NEUN- and OLIG2+/NEUN-), pt9 (none), pt12 (PU.1+/NEUN-), and pt14 (PU.1+/NEUN-).

Article Snippet: Nuclei were resuspended in staining buffer (2% bovine serum albumin, 1 mM EDTA and phosphate-buffered saline) and immunostained overnight at 4 °C with the following primary antibodies: conjugated anti-NEUN-PE for neurons (1:1,000, Milli-Mark, cat. no. FCMAB317PE) , conjugated anti-PU.1-AF647 for microglia (1:100, Cell Signaling Technology, cat. no. 2240S conjugate) , , unconjugated anti-OLIG2 for oligodendrocytes (1:500, Abcam, cat. no. ab109186) , conjugated anti-PAX6-APC for astrocytes (1:1,000, Novus Biologicals, cat. no. NBP2-34705APC) and rabbit unconjugated anti-TBR1 for excitatory neurons (1:1,000, Abcam, cat. no. ab31940) .

Techniques: Expressing, Mutagenesis, Marker, Fluorescence, Selection, Staining, Amplification, Sequencing, Variant Assay

Journal: Nature Neuroscience

Article Title: Single-cell genotyping and transcriptomic profiling of mosaic focal cortical dysplasia

doi: 10.1038/s41593-025-01936-z

Figure Lengend Snippet: a-e , Validation of transcriptional changes (that is genes with absolute log 2 (FC) > 0.3) between mutation detected (Mut.) and reference detected (Ref.) glutamatergic neurons (GluN) and astrocytes (Astro). Box plots depict the median and interquartile range, with whiskers indicating minimum and maximum values. a-b , Differential expression analysis using 50 random subset comparisons of n = 29 GluN ( a ) or n = 17 astrocytes ( b ) amongst Mut. and Ref. nuclei of pt9 and pt10 compared to the rest of GluN and astrocytes, respectively. Left, proportion of shared dysregulated genes in ‘random’ vs ‘observed’ Mut. vs Ref. comparisons. Right, Jaccard similarity index. Wilcoxon signed rank test with continuity correction confirms significant differences between ‘random’ and ‘observed’ dysregulated genes in both GluN and astrocytes. c , Linear regression model using ‘random’ dysregulated genes from iteration n.1 of GluN and Astro cannot predict Mut. vs Ref. nuclei. Box plots depict the median and interquartile range, with whiskers indicating minimum and maximum values. d , Linear regression model using observed dysregulated genes successfully discriminates Mut. from Ref. nuclei for both GluN and Astro, with potential false negatives identified in 14% (1/7) of pt9 and 9% (8/85) of pt10 Ref. GluN nuclei (indicated by black dotted box above red threshold line). Box plots depict the median and interquartile range, with whiskers indicating minimum and maximum values. e , Expression patterns of GluN and Astro ‘observed’ dysregulated genes across patients (pt9-10) and controls (ct1-3). Analysis restricted to samples with >10 Mut./Ref. nuclei. Statistical analysis of average gene expression in Mut. vs. Ref. GluN by individual using Kruskal-Wallis test shows significant mutation effects (genes upregulated in Mut.: H statistics = 340.50, FDR-adjusted p-value: 9.92×10 − 76; genes downregulated in Mut.: H statistics = 30.59, FDR-adjusted p-value: 3.19×10 − 8). No statistical test was performed for astrocytes since only one patient with > 10 Mut. nuclei was available. f , Top 10 GO terms for Mut. vs. Ref. dysregulated genes in astrocytes. g , Mean expression of GluN dysregulated genes between Ref. nuclei from patients vs. controls in Mut., Ref. and control GluN from pt9-10 and ct1-2. Left, all dysregulated genes. Right, epilepsy-related dysregulated genes. h , Top 10 GO terms for patient Ref. vs. control. nuclei dysregulated genes. Only significant GO terms (adjusted p-value < 0.05) are shown.

Article Snippet: Nuclei were resuspended in staining buffer (2% bovine serum albumin, 1 mM EDTA and phosphate-buffered saline) and immunostained overnight at 4 °C with the following primary antibodies: conjugated anti-NEUN-PE for neurons (1:1,000, Milli-Mark, cat. no. FCMAB317PE) , conjugated anti-PU.1-AF647 for microglia (1:100, Cell Signaling Technology, cat. no. 2240S conjugate) , , unconjugated anti-OLIG2 for oligodendrocytes (1:500, Abcam, cat. no. ab109186) , conjugated anti-PAX6-APC for astrocytes (1:1,000, Novus Biologicals, cat. no. NBP2-34705APC) and rabbit unconjugated anti-TBR1 for excitatory neurons (1:1,000, Abcam, cat. no. ab31940) .

Techniques: Biomarker Discovery, Mutagenesis, Quantitative Proteomics, Expressing, Gene Expression, Control

Journal: Nature Neuroscience

Article Title: Single-cell genotyping and transcriptomic profiling of mosaic focal cortical dysplasia

doi: 10.1038/s41593-025-01936-z

Figure Lengend Snippet: a , Left, LCM–seq workflow for capturing pools of DNs, BCs and NNs from eight patients (pt1–5 and pt7–9). Right, heatmap of NEFM and VIM normalized expression with unsupervised hierarchical clustering. b , Label transfer of LCM–seq samples on to the snRNA-seq UMAP space showing NNs or DNs matching with GluNs and BCs with astrocytes. c , Left, NRGN and GFAP normalized expression heatmap with unsupervised hierarchical clustering. Right, co-immunofluorescence showing NRGN in pS6 + /SMI311 + DNs and GFAP in pS6 + /VIM + BCs (pt5) ( n = 1 section/patient/staining analyzed). GFAP-pS6 and VIM-pS6 double stainings were performed on two consecutive sections and the same BC was recognized in both sections. Nuclei (in blue) are labeled with DAPI. Scale bars, 50 µm. d , Visium spatial transcriptomics showing intermingled spots containing DNs and BCs across the tissue (pt5). Magnified images show representative DN- and BC-containing spots after hematoxylin and eosin staining ( n = 1 section per patient analyzed). Scale bars, 1.5 mm; insets = 55 µm. e , Top markers of DN- and BC-containing spots (pt5). Known histological markers for DNs ( NEFM ) and BCs ( CRYAB ) are enriched in spots with DNs and BCs. f , Spatial semi-supervised clustering of Visium spots showing clusters enriched in GluNs, astrocytes and oligodendrocytes (pt5) with top marker genes in parentheses. g , Distinct clusters for DNs, BCs, astrocytes (Astros) and GluNs from single cells (pt5 and pt9) of the MERSCOPE UMAP space. h , Heatmap of the top ten DN or BC markers with representative MERSCOPE images (pt5). DNs are identified as pS6 + /NEUN + and BCs as pS6 + /NEUN − ( n = 1 section per patient analyzed). Scale bars, 50 µm. i , Left, number of shared dysregulated genes across Mut. versus Ref. GluNs (snRNA-seq), DNs versus NNs (LCM–seq) and DN-containing spots (Visium). Right, top GO terms of DN upregulated genes. Ribo-nt., ribonucleotides; metab., metabolic; proc., process; Ribo-ns., ribonucleosides; RP., ribosomal proteins; rNTP, ribonucleoside triphosphates. j , Representative images of strong VDAC1 immunostaining in pS6 + DNs (pt2) ( n = 1 section/patient/staining analyzed). Scale bars, 50 µm. k , Electron microscopy of DNs (pt5) showing an accumulation of vesicular, swollen, damaged mitochondria (black circles) ( n = 1 section per patient analyzed). Scale bar, 2.5 µm. Detailed sample information for each experiment and analysis is provided in Supplementary Table . expr., expression; max., maximum; min., minimum.

Article Snippet: Nuclei were resuspended in staining buffer (2% bovine serum albumin, 1 mM EDTA and phosphate-buffered saline) and immunostained overnight at 4 °C with the following primary antibodies: conjugated anti-NEUN-PE for neurons (1:1,000, Milli-Mark, cat. no. FCMAB317PE) , conjugated anti-PU.1-AF647 for microglia (1:100, Cell Signaling Technology, cat. no. 2240S conjugate) , , unconjugated anti-OLIG2 for oligodendrocytes (1:500, Abcam, cat. no. ab109186) , conjugated anti-PAX6-APC for astrocytes (1:1,000, Novus Biologicals, cat. no. NBP2-34705APC) and rabbit unconjugated anti-TBR1 for excitatory neurons (1:1,000, Abcam, cat. no. ab31940) .

Techniques: Expressing, Immunofluorescence, Staining, Labeling, Marker, Immunostaining, Electron Microscopy

Journal: Nature Neuroscience

Article Title: Single-cell genotyping and transcriptomic profiling of mosaic focal cortical dysplasia

doi: 10.1038/s41593-025-01936-z

Figure Lengend Snippet: a , Semi-supervised clustering of Visium data from patients pt2, pt5 and pt9. Clusters (highlighted in red) are annotated based on predominant marker genes. To differentiate clusters enriched for the same cell type, the top expressed marker was added to the cluster annotation. Clusters lacking clear cell-type enrichment in pt9 are labeled ‘Unknown’. Abbreviations: N., neurons; IN, interneurons; NF high, cluster with high levels of neurofilament genes; Astro, astrocytes; Oligo, oligodendrocytes; Micro, microglia. On the right: corresponding hematoxylin and eosin (HE) -stained section. Scale bars = 1.5 mm. b , Spatial distribution of DN/BC-containing spots in pt2 and pt9 showing distribution across both anatomical space and transcriptionally-defined clusters. To differentiate clusters enriched for the same cell type, the top discriminating marker expressed was added to the cluster annotation. Scale bars = 1.5 mm. c , Expression analysis of cluster-defining markers in each Visium sample. Dot size indicates percentage of expressing spots; color intensity shows average normalized expression. Abbreviations: Pct. Expr., percentage of spots expressing the genes; Avg. Exprs., average normalized gene expression per group. d , Spatial distribution of cortical layer marker expression scores calculated using previously defined gene sets (Maynard et al., 2021) . Higher scores indicate increased expression of the gene set.

Article Snippet: Nuclei were resuspended in staining buffer (2% bovine serum albumin, 1 mM EDTA and phosphate-buffered saline) and immunostained overnight at 4 °C with the following primary antibodies: conjugated anti-NEUN-PE for neurons (1:1,000, Milli-Mark, cat. no. FCMAB317PE) , conjugated anti-PU.1-AF647 for microglia (1:100, Cell Signaling Technology, cat. no. 2240S conjugate) , , unconjugated anti-OLIG2 for oligodendrocytes (1:500, Abcam, cat. no. ab109186) , conjugated anti-PAX6-APC for astrocytes (1:1,000, Novus Biologicals, cat. no. NBP2-34705APC) and rabbit unconjugated anti-TBR1 for excitatory neurons (1:1,000, Abcam, cat. no. ab31940) .

Techniques: Marker, Labeling, Staining, Expressing, Gene Expression

Journal: Molecular Vision

Article Title: Embryonic stem cell–derived photoreceptor precursor cells differentiated by coculture with RPE cells

doi:

Figure Lengend Snippet: Primary antibody information.

Article Snippet: PAX6 , 1:100 , Novus biologicals , , Littleton, CO.

Techniques: