nanog primary  (R&D Systems)

Journal: Scientific reports

Article Title: Topological data analysis of pattern formation of human induced pluripotent stem cell colonies.

doi: 10.1038/s41598-025-90592-1

Figure Lengend Snippet: Fig. 2. Artificial induction of exogenous GATA6-HA occurs within the context of endogenous GATA6 expression. (a) Gene circuit for chemical induction of GATA6-HA expression. The pan-GATA6 antibody can detect both induced GATA6-HA and endogenous GATA6 (i.e. the total amount of GATA6), whereas the HA antibody can only detect induced GATA6-HA (i.e. a subset produced via induction of the total amount of GATA6). (b) Representative immunofluorescence images of NANOG and pan-GATA6 or HA at 0 and 25 ng/ml Dox concentrations (scale bar, 440µm). Using Volocity, we applied the gamma changes (gamma of 1.5) after brightness enhancement on all stitched large images used in the figure for better contrast for representation (see Methods). (c) Quantification of segmented images by each channel from (b). Green (NANOG) and red (pan-GATA6 or HA) fluorescent intensities are normalized to the corresponding nuclear Hoechst value in blue. The threshold for the signal in the green channel is given by the green dotted line, and the red dotted line indicates the threshold for the signal in the red channel. The four cell types based on these thresholds are given by the labels R+G−, R+G+, R−G+, and R−G− of the corresponding quadrant.

Article Snippet: Primary antibodies are Nanog (Cell Signaling Technology 4893S, 1:2000), Gata6 (Abcam ab22600, 1:200, R&D Systems AF1700, which was only used in co-staining, 1:200), and HA (Novus Biologicals NB600-363R, 1:200).

Techniques: Expressing, Produced, Immunofluorescence

Journal: Stem Cells

Article Title: NK-cell cytotoxicity toward pluripotent stem cells and their neural progeny: impacts of activating and inhibitory receptors and KIR/HLA mismatch

doi: 10.1093/stmcls/sxae083

Figure Lengend Snippet: B7H6 expression is maintained in motoneurons differentiated from iPS-derived neural progenitor cells. (A) B7H6 (NCR3LG1) transcription was assessed by qPCR analysis using cDNA from the indicated cells, displaying relative expression using GNB2LI as housekeeping control. The tumor cell lines HCT15 and MDA.MB.231 have been included as B7H6+ and B7H6− controls, respectively. The results are normalized and represent 3 or more independent triplicate experiments. Neurons were generated from iPS-derived NPCs, and their phenotype was documented by (B) phase contrast microscopy demonstrating dendritic morphology, and (C) qPCR analysis for the neuronal markers NeuN (RBFOX3), MAP-2 (MAP2), class III β-tubulin (TuJ1; gene TUBB3), and synaptotagmin (SYT1), as indicated. NPC phenotype was defined by downregulation of the pluripotency marker Oct4 (POU5F1) and concomitant upregulation of nestin (NES). Expression was calculated using β-actin as an internal control. Error bars indicate SD based on 1 triplicate experiment.

Article Snippet: Following permeabilization in 0.1% Triton, the cells were stained with primary antibodies against the pluripotency markers Nanog (clone 7F7.1; 1:400; Millipore), Oct4, and SOX2 (cat. no. 090023 and 090024, respectively; 1:100; StemGent) overnight at 4 °C, washed, incubated for 1 hour at room temperature with Cy2-conjugated or Cy3-conjugated secondary antibodies (1:400; Jackson ImmunoResearch) followed by counterstaining of nuclei with DAPI.

Techniques: Expressing, Derivative Assay, Control, Generated, Microscopy, Marker