anti jagged 1  (Novus Biologicals)

Journal: Nature Communications

Article Title: Controlling nephron precursor differentiation to generate proximal-biased kidney organoids with emerging maturity

doi: 10.1038/s41467-025-63107-9

Figure Lengend Snippet: a UMAP of single-cell RNA-sequencing from day 10 (pre-treatment), day 12 control, and day 12 PI3K inhibitor (LY294002)-treated kidney organoid nephrons. Cells are colored by sample with marker gene annotations. b Heatmap of Z-scores for hierarchically clustered differentially expressed genes across samples. Representative genes are listed, with a Z-score legend. c Feature plots of genes enriched in each sample group from the differentially expressed gene list. d Feature plots of selected genes for each group based on sample characteristics. For c , d human nephron single-cell detection (Supplementary Fig. ) is shown on the left, organoid plot on the right. e UMAP of single-cell RNA-sequencing from day 12 control and day 12 LY294002-treated kidney organoid nephrons. Cells match a but exclude day 10 sample. Cells are colored by sample with marker gene annotations. f Heatmap of log 2 fold change (log 2 FC) for differentially expressed genes, calculated as the ratio of LY294002-treated to control aggregate expression. Representative genes are listed with a log 2 FC legend. g Feature plots of genes enriched in each sample from the differentially expressed gene list. Human nephron single-cell (Supplementary Fig. ) detection is on the left, organoid plot on the right. h Split violin plots of genes from the differentially expressed gene list. Top two rows show genes enriched in day 12 control cells; bottom two rows show genes enriched in day 12 LY294002-treated cells. Plot colors match sample colors from ( e ). i Gene ontology terms for the top 50 differentially expressed genes per sample. Top graph: day 12 control nephron cells; bottom graph: LY294002-treated nephron cells. j–l Whole-mount immunofluorescent stains of day 12 control and LY294002-treated kidney organoids. Boxed regions are magnified. Inset in j shows absence of detectable HNF4A protein (orange) in day 12 samples. Scale bars: 10 microns. m Whole-mount immunofluorescent stain of day 12 control and LY294002-treated kidney organoids. Insets highlight JAG1 and HNF1B protein detection. Scale bars: 500 microns. n Quantification of JAG1 + and HNF1B + nephron size (µm 2 ) and HNF1B + intensity (RFU) for n = 3 day 12 organoids each, across all positive segments. SEM error bars shown. Statistical significance determined by two-sided Student’s t -test.

Article Snippet: Primary antibodies used in this study were: WT1 (abcam, ab89901, 1:1000), JAG1 (R&D Systems, AF599, 1:300), HNF1B (Thermo Fisher Scientific, MA5-24605, 1:500), HNF4A (R&D Systems, MAB4605, 1:200), CDH1 (BD Biosciences, 610181, 1:300), ZO-1 (Thermo Fisher Scientific, 33-9100, 1:200), PAX2 (R&D Systems, AF3364, 1:50), SIX1 (Cell Signaling Technology, 12891S, 1:300), HES1 (Cell Signaling Technology, 11988, 1:300), POU3F3 (Novus Biologicals, NBP1-49872, 1:500), HNF4G (Thermo Fisher Scientific, PA5-82189, 1:200), LRP2 (My Bio Source, MBS690201, 1:500), HAVCR1 (R&D Systems, AF1750, 1:200), γH2AX (Cell Signaling Technology, 2577), LAMB1 (Santa Cruz Biotechnology, sc-33709, 1:250), ATP1A1 (Abcam, ab7671, 1:200), SOX9 (Abcam, ab185230, 1:300), PODXL (R&D Systems, AF1658, 1:200), PAX8 (Abcam, ab189249, 1:100), LEF1 (Santa Cruz Biotechnology, sc-374412, 1:200), LTL (Vector Laboratories, B-1325-2, 1:300), SLC12A1 (Sigma Aldrich, HPA018107, 1:200), TFAP2A (Santa Cruz Biotechnology, sc-12726, 1:200), Alexa 647-conjugated LRP2 (R&D Systems, FAB9578R, 1:100), and Alexa 594-conjugated LRP2 (R&D Systems, FAB9578T, 1:100).

Techniques: RNA Sequencing, Control, Marker, Expressing, Staining