Journal:

Article Title: Human airway epithelial cell determinants of survival and functional phenotype for primary human mast cells

doi: 10.1073/pnas.0503948102

Figure Lengend Snippet: MC protease expression identified by immunostaining over time. MCs were recovered after 2 d of culture in liquid suspension culture with SCF alone, in direct coculture with HAECs, in HAEC coculture separated by a transwell membrane, with HAEC-conditioned

Article Snippet: Analysis of SCF protein expression from HAEC supernatants was performed by using a sandwich ELISA specific for human SCF (Chemicon).

Techniques: Expressing, Immunostaining

Journal:

Article Title: Human airway epithelial cell determinants of survival and functional phenotype for primary human mast cells

doi: 10.1073/pnas.0503948102

Figure Lengend Snippet: SCF protein expression by HAECs. Release of soluble SCF protein into HAEC culture media was detected by SCF-ELISA. ( A ) All data are mean ± SEM ( n = 4). Detection of membrane-bound SCF expression by immunohistochemical staining of HAECs by using

Article Snippet: Analysis of SCF protein expression from HAEC supernatants was performed by using a sandwich ELISA specific for human SCF (Chemicon).

Techniques: Expressing, Enzyme-linked Immunosorbent Assay, Immunohistochemistry, Staining

Journal:

Article Title: Human airway epithelial cell determinants of survival and functional phenotype for primary human mast cells

doi: 10.1073/pnas.0503948102

Figure Lengend Snippet: Expression of SCF mRNA by HAECs. Soluble (572 bp) and membrane-bound (488 bp) SCF expression by HAECs was identified by RT-PCR analysis. SCF cDNA was amplified by 35 cycles of PCR. Lanes 1 and 2, HAEC donor 1; lanes 3 and 4, primary human MC; lanes 5

Article Snippet: Analysis of SCF protein expression from HAEC supernatants was performed by using a sandwich ELISA specific for human SCF (Chemicon).

Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, Amplification, Polymerase Chain Reaction

Journal: Journal of cellular physiology

Article Title: OSCILLATING FLUID FLOW ACTIVATION OF GAP JUNCTION HEMICHANNELS INDUCES ATP RELEASE FROM MLO-Y4 OSTEOCYTES

doi: 10.1002/jcp.21021

Figure Lengend Snippet: Cx43 siRNA decreases Cx43 protein expression and inhibits oscillatory fluid flow-induced dye uptake. (A) Western blotting of Cx43 protein levels demonstrated a reduction in Cx43 expression in MLO-Y4 cells treated with Cx43 siRNA, but not scrambled, non-silencing

Article Snippet: Cx43 protein expression in scrambled siRNA-transfected, and Cx43 siRNA-transfected cells was examined with a monoclonal anti-Cx43 antibody from Chemicon and visualized using enhanced chemiluminescent detection (Amersham Biosciences); membranes were then stripped and re-probed for GAPDH (Accurate Scientific & Chemical Corp.) to confirm equal loading of all lanes.

Techniques: Expressing, Flow Cytometry, Western Blot

Journal: Journal of cellular physiology

Article Title: OSCILLATING FLUID FLOW ACTIVATION OF GAP JUNCTION HEMICHANNELS INDUCES ATP RELEASE FROM MLO-Y4 OSTEOCYTES

doi: 10.1002/jcp.21021

Figure Lengend Snippet: ATP release in MLO-Y4 osteocytes in response to oscillatory fluid flow requires hemichannel activation and Cx43 expression. (A) Oscillating fluid flow significantly increased ATP content in conditioned media relative to static controls. Data were obtained

Article Snippet: Cx43 protein expression in scrambled siRNA-transfected, and Cx43 siRNA-transfected cells was examined with a monoclonal anti-Cx43 antibody from Chemicon and visualized using enhanced chemiluminescent detection (Amersham Biosciences); membranes were then stripped and re-probed for GAPDH (Accurate Scientific & Chemical Corp.) to confirm equal loading of all lanes.

Techniques: Flow Cytometry, Activation Assay, Expressing

Journal: Scientific Reports

Article Title: Early embryonic exposure of freshwater gastropods to pharmaceutical 5-alpha-reductase inhibitors results in a surprising open-coiled “banana-shaped” shell

doi: 10.1038/s41598-019-52850-x

Figure Lengend Snippet: B. glabrata embryo lysate reacts with both 5αR1 and 5αR2 antibodies in Western blot assays, ( a ) bands can be seen for both SRD5A1 transfected 293 T cell lysate (Abnova, 10 µl) in lane 3 and B. glabrata embryo lysates in lanes 1 2 (30 µg protein per lane), ( b ) B. glabrata embryo lysate produces a strong reaction with human 5αR2 antibodies in lane 3 (30 µg protein per lane), human PC3 prostate cells lysate (included as a ‘positive’ response marker) is also visible but fainter in lanes 1 2 (170 µg protein per lane). The total protein content of both snail embryo and PC3 cell culture lysates were quantified using the Bradford assay. For both blots 10 µg of protein ladder was used per lane, (blot a, 10 µg Color Prestained Protein Standard, New England Bio-Labs, and blot b, 10 µg Precision Plus Protein™ Dual Colour Standards, Bio-rad). Images of gels have been cropped to focus on the sample bands. In each gel, only 3 samples (as described) were run. Blots for reactivity with SRD5A1 and SRD5A2 were run on separate gels and occasions. Images captured of full-length gels with additional exposures can be seen in Supporting Information Figs S1 and S2 .

Article Snippet: 5α-reductase proteins in B. glabrata embryos To detect 5αR1 and 5αR2 protein expression in B. glabrata embryos, replicates of 100 hippo stage embryos (4-days post-fertilisation) were dissected from their egg masses and homogenised in 100 µL of cold RIPA buffer (Sigma Aldrich, UK) with a cocktail of protease inhibitor (Roche).

Techniques: Western Blot, Transfection, Marker, Cell Culture, Bradford Assay

Journal: Communications Biology

Article Title: The EphA2 receptor is activated through induction of distinct, ligand-dependent oligomeric structures

doi: 10.1038/s42003-018-0017-7

Figure Lengend Snippet: EphA2 dimerization induced by the monomeric ephrinA1 ligand. a MSE vs. oligomer order for EphA2 wild-type and the L223R/L254R/V255R and G131Y mutants in the presence of 200 nM m-ephrinA1. The MSEs are all minimized for n = 2, indicating dimerization. b Comparison of EphA2 dimerization propensity in the presence and absence of m-ephrinA1 shows that m-ephrinA1 significantly enhances EphA2 dimerization. c Dimerization curves in the presence of m-ephrinA1 show that the dimerization propensity of EphA2 wild-type and the L223R/L254R/V255R mutant are the same, while the G131Y mutant has a reduced dimerization propensity, indicating the involvement of the dimerization interface. d A representative Western blot showing Y772 phosphorylation of EphA2 wild-type and the indicated mutants following a 15 min stimulation with m-ephrinA1 and FBS. e Quantification of Y772 phosphorylation from three to four independent measurements is shown as solid circles. The bars represent the averages and the standard errors. EphA2 wild-type and the L223R/L254R/V255R mutant exhibit similar levels of Y772 phosphorylation while the G131Y mutant shows significantly lower phosphorylation (** p

Article Snippet: EphA2 expression was quantified using anti-EphA2 antibodies (Cell Signalling, MA).

Techniques: Mutagenesis, Western Blot

Journal: Communications Biology

Article Title: The EphA2 receptor is activated through induction of distinct, ligand-dependent oligomeric structures

doi: 10.1038/s42003-018-0017-7

Figure Lengend Snippet: Cartoon representation of the findings: EphA2 can associate into two types of dimers as well as clusters, depending of the nature of the activating ligand. TK tyrosine kinase, FP fluorescent protein

Article Snippet: EphA2 expression was quantified using anti-EphA2 antibodies (Cell Signalling, MA).

Techniques:

Journal: Communications Biology

Article Title: The EphA2 receptor is activated through induction of distinct, ligand-dependent oligomeric structures

doi: 10.1038/s42003-018-0017-7

Figure Lengend Snippet: Effect of the R103E mutation on EphA2 dimerization in the presence of monomeric ephrinA1 ligand. a MSE vs. oligomer order for the EphA2 R103E mutant in the presence of m-ephrinA1. The MSE is miminized for n = 2, indicating dimerization. b Dimerization propensity of the EphA2 R103E mutant in the absence and in the presence of m-ephrinA1. The ligand slightly enhances dimerization, indicating that the EphA2 R103E mutation does not completely abrogate m-ephrinA1 binding. c Dimerization curves for EphA2 wild-type and the R103E mutant in the presence of m-ephrinA1. The R103E mutation decreases the stability of the dimers, most likely because it severely impairs m-ephrinA1 binding

Article Snippet: EphA2 expression was quantified using anti-EphA2 antibodies (Cell Signalling, MA).

Techniques: Mutagenesis, Binding Assay

Journal: Communications Biology

Article Title: The EphA2 receptor is activated through induction of distinct, ligand-dependent oligomeric structures

doi: 10.1038/s42003-018-0017-7

Figure Lengend Snippet: EphA2 dimerization in the presence of the monomeric YSA peptide ligand. a MSE vs. oligomer order for EphA2 wild-type and the L223R/L254R/V255R, G131Y, and R103E mutants. In all cases, the MSE minimum occurs at n = 2, indicating dimerization. b Dimerization curves show that the dimerization propensity of EphA2 wild-type and the G131Y mutant are the same, while the L223R/L254R/V255R mutant has reduced dimerization propensity. Data for the wild-type and the L223R/L254R/V255R mutant are from ref. 30 . c Dimerization curves for EphA2 wild-type and the R103E mutant show that the mutant has reduced dimerization propensity

Article Snippet: EphA2 expression was quantified using anti-EphA2 antibodies (Cell Signalling, MA).

Techniques: Mutagenesis

Journal: Communications Biology

Article Title: The EphA2 receptor is activated through induction of distinct, ligand-dependent oligomeric structures

doi: 10.1038/s42003-018-0017-7

Figure Lengend Snippet: Dimeric ephrinA1-Fc induces EphA2 receptor clustering. a Portion of a HEK293T cell expressing EphA2-mTurq and EphA2-eYFP in hypo-osmotic medium, imaged when mTurq was excited. A plasma membrane region of homogeneous fluorescence, a few µm in length (yellow box), is analyzed to determine the EphA2-mTurq concentration, the EphA2-eYFP concentration and the FRET efficiency, as described in the Materials and methods. The scale bar is 5 μm. b FRET efficiency vs. acceptor (EphA2-eYFP) concentration. Each data point corresponds to one plasma membrane region. c Donor (EphA2-mTurq) concentration vs. acceptor (EphA2-eYFP) concentration in the selected membrane regions. In b and c , 275 cells were imaged in four independent experiments, yielding 858 data points. d Mean square error (MSE) vs. oligomer order. MSE is minimized for n > 4, indicating the presence of oligomers that are larger than dimers (i.e., clusters). e Clustered EphA2 receptor fraction as a function of total EphA2 concentration. The data were binned and the averages are shown along with the standard errors. The solid line represents the theoretical best fit to the data. f Mean FRET efficiencies vs. mean acceptor fractions, determined as shown in Supplementary Figure 1 . The plot is based on more than 1000 data points. The dependence deviates from a linear function, supporting the conclusion that exposure to ephrinA1-Fc induces preferentially the formation of EphA2 clusters

Article Snippet: EphA2 expression was quantified using anti-EphA2 antibodies (Cell Signalling, MA).

Techniques: Expressing, Fluorescence, Concentration Assay

Journal: Communications Biology

Article Title: The EphA2 receptor is activated through induction of distinct, ligand-dependent oligomeric structures

doi: 10.1038/s42003-018-0017-7

Figure Lengend Snippet: Interfaces involved in EphA2 receptor clustering induced by ephrinA1-Fc. a Crystal structure showing a lateral view of four EphA2 extracellular regions (gray) bound to four ephrinA1 molecules (light blue; PDB ID: 3MX0). The receptor tetramer is stabilized via two interfaces: the “clustering” interface (approximately outlined in orange), which includes contacts mediated by L223, L254, and V255 in the cysteine-rich domain, and the dimerization interface (approximately outlined in wine), which includes contacts mediated by G131 in the ligand-binding domain 18 . b Comparison of raw FRET data for EphA2 wild-type and the L223R/L254R/V255R mutant in the presence of ephrinA1-Fc. In this experiment, 275 cells were imaged in four independent experiments to obtain 858 data points for the wild-type, and 196 cells were imaged in four independent experiments to obtain 563 data points for the L223R/L254R/V255R mutant. c Comparison of raw FRET data for EphA2 wild-type and the G131Y mutant in the presence of ephrinA1-Fc. A total of 618 cells were imaged in six independent experiments to yield 2310 data points for the G131Y mutant. d MSE vs. oligomer order for the L223R/L254R/V255R and G131Y mutants in the presence of ephrinA1-Fc. The MSE minimum for the L223R/L254R/V255R mutant occurs at n = 6. The MSE for the G131Y mutant is the same for n ≥ 2. As previously shown 21 , these results indicate that the EphA2 receptor is preferentially assembled into clusters, although the presence of some dimers cannot be excluded. e Representation of the clustered fractions for EphA2 wild-type and the L223R/L254R/V255R and G131Y mutants as a function of total receptor concentration shows that mutation of both interfaces decreases the fraction of clustered EphA2

Article Snippet: EphA2 expression was quantified using anti-EphA2 antibodies (Cell Signalling, MA).

Techniques: Ligand Binding Assay, Mutagenesis, Concentration Assay

Journal: Communications Biology

Article Title: The EphA2 receptor is activated through induction of distinct, ligand-dependent oligomeric structures

doi: 10.1038/s42003-018-0017-7

Figure Lengend Snippet: Dimerization of EphA2 wild-type and the L223R/L254R/V255R, G131Y and R103E mutants in the absence of ligand binding. a MSE vs. oligomer order for EphA2 wild-type and the three mutants. MSE is minimized at n = 2 for all, indicating the presence of dimers. b Dimerization curves for EphA2 wild-type and the L223R/L254R/V255R and G131E mutants. The L223R/L254R/V255R mutations reduce dimerization, while the G131Y mutation has no effect. Thus, the unliganded dimer is stabilized through the “clustering interface”. c Dimerization curves for EphA2 wild-type and the R103E mutant. The R103E mutant exhibits a reduced dimerization propensity, despite the fact that this residue is not part of the clustering interface. d A representative Western blot comparing Y772 phosphorylation for EphA2 wild-type and the G131Y mutant. e Quantification from three independent experiments (shown as solid circles) shows no statistically significant difference ( p > 0.05 from Student’s t -test). f A representative Western blot image comparing S897 phosphorylation for EphA2 wild-type and the G131Y mutant. g Quantification from four independent experiments (shown as solid circles) shows no statistically significant difference ( p > 0.05 from Student’s t -test). h Representative Western blot images comparing Y772 phosphorylation for EphA2 wild-type and the R103E mutant. i Quantification from three independent experiments (shown as solid circles) shows that the R103E mutant has lower Y772 phosphorylation than EphA2 wild-type (*** p

Article Snippet: EphA2 expression was quantified using anti-EphA2 antibodies (Cell Signalling, MA).

Techniques: Ligand Binding Assay, Mutagenesis, Western Blot

Journal: Communications Biology

Article Title: The EphA2 receptor is activated through induction of distinct, ligand-dependent oligomeric structures

doi: 10.1038/s42003-018-0017-7

Figure Lengend Snippet: Effect of the R103E mutation on EphA2 clustering induced by ephrinA1-Fc. a , b Side and top views of a crystal structure of four EphA2 molecules (gray) bound to four ephrinA1 molecules (light blue; PDB ID: 3MX0). The position of the R103E mutation in two of the EphA2 molecules is shown in magenta, and indicated by an arrow in one. c Comparison of raw FRET data for EphA2 wild-type and the R103E mutant in the presence of ephrinA1-Fc. 275 cells were imaged in four independent experiments to obtain 858 data points for the wild type. A total of 201 cells were imaged in three independent experiments to obtain 474 data points for R03E mutant. d MSE vs. oligomer order for EphA2 R103E in the presence of ephrinA1-Fc. The MSE value is the same for all n ≥ 2, indicating oligomerization with predominance of clusters. e Comparison of EphA2 wild-type and R103E mutant clustered fractions in the presence of saturating concentration of ephrinA1-Fc shows that the R103E mutation severely destabilizes the clusters

Article Snippet: EphA2 expression was quantified using anti-EphA2 antibodies (Cell Signalling, MA).

Techniques: Mutagenesis, Concentration Assay

Journal: Frontiers in Cellular Neuroscience

Article Title: MAP Kinase Phosphatase 3 (MKP3) Preserves Norepinephrine Transporter Activity by Modulating ERK1/2 Kinase-Mediated Gene Expression

doi: 10.3389/fncel.2017.00253

Figure Lengend Snippet: Knockdown of caveolin-1 attenuates PKC-mediated down regulation of NET function. Immunoblotting demonstrate silencing of caveolin-1 expression by stable expression of Cav1 shRNA construct (PC12-Cav1 shRNA) or MKP3 (PC12-MKP3) and successful re-introduction of caveolin-1 in MKP3 expressing cells (PC12-MKP3-HACav1) (A) . Following Cav1 silencing or re-introduction PKC was activated by PMA for 30 min and NET-specific radiolabeled uptake was performed as above (B) . Results are displayed as percentage uptake following PMA treatment compared to vehicle treated control of respective cell type. Expression of MKP3 or silencing of Caveolin1 by the shRNA construct attenuates NET downregulation following PKC activation to a similar degree (B) . Re-introduction of caveolin1 into MKP3 expressing PC12 cells does not recover the PKC-mediated down regulation (B) . All experiments were performed in triplicate in three independent experiments. Results from three independent experiments were analyzed by two-way ANOVA with Bonferroni post hoc analysis (** p

Article Snippet: To detect levels of caveolin-1 expression antibodies from Santa Cruz Biotechnology was used (N-20, sc-894).

Techniques: Expressing, shRNA, Construct, Activation Assay

Journal: Scientific Reports

Article Title: CMV promoter mutants with a reduced propensity to productivity loss in CHO cells

doi: 10.1038/srep16952

Figure Lengend Snippet: ( a ) hCMV-MIE sequence (left) and list of mutants (right). Positions where C to G mutations were introduced are marked with an asterisk. Transcription factor binding sites are underlined. CpG sites are highlighted in grey. hCMV-MIE variants are coded with a three letter identifier referring to the nucleotides at positions −508, −179 and −41. DNA positions are numbered relative to the transcription start site. (b) Transient expression of SEAP driven by hCMV-MIE mutants. Expression levels were normalised to unmutated hCMV-MIE (CCC). Error bars represent SD of eight biological replicates. Two independent experiments were performed. Potential effects of the hCMV-MIE mutations within the same experiment were tested with Tukey HSD (α = 0.05). Significant differences between the promoter mutants and CCC within the same experiment are marked with the respective p-value. (c) eGFP expression of permanently transfected CHO cell pools 34 days (upper panel) and 87 days (lower panel) after transfection. eGFP expression was quantified by FACS and the geometrical mean of the fluorescence intensity (gMFI) was plotted for each cell pool. The identity of the promoter variants and the number (N) of independent cell pools is indicated below the x-axis. The upper and the lower ends of the boxes represent the first and the third quartile of each group. The ends of the whiskers represent the lowest and highest values still within the 1.5fold interquartile range. The grey line indicates the overall mean of the gMFI values. (d) eGFP expression of permanently transfected CHO cell pools 42 days (left panel) and 69 days (right panel) after transfection; all further labelling is identical to ( c ). (e) Percentage methylation levels of hCMV-MIE variants at each CpG cytosine 42 days (upper panels) and 69 days (lower panels) after transfection. Methylation levels are plotted against DNA positions numbered 5′ to 3′. The positions of C-508, C-179, and C-41 are highlighted.

Article Snippet: Analysis of eGFP and antibody expression eGFP expression in CHO-K1 was examined before passaging the cells with a BD FACS Canto II flow cytometer (BD, Heidelberg, Germany).

Techniques: Sequencing, Binding Assay, Expressing, Countercurrent Chromatography, Transfection, FACS, Fluorescence, Methylation

Journal:

Article Title: Regulation of insulin secretion and GLUT4 tra3cking by the calcium sensor synaptotagmin VII ☆

doi: 10.1016/j.bbrc.2007.08.023

Figure Lengend Snippet: Dysregulated GLUT4 traffic in Syt VII−/− mice. (A) GLUT4 traffic in WT and Syt VII−/− fat cells: GLUT4 distribution examined by confocal microscopy in adipocytes, isolated from wild-type littermate (WT, n = 3) and Syt VII−/−,

Article Snippet: GLUT4 expression was detected in adipocytes by immunofluorescence (anti-GLUT4 antibody, Cell Signaling Technology, Beverly, MA).

Techniques: Mouse Assay, Confocal Microscopy, Isolation

Journal:

Article Title: Regulation of insulin secretion and GLUT4 tra3cking by the calcium sensor synaptotagmin VII ☆

doi: 10.1016/j.bbrc.2007.08.023

Figure Lengend Snippet: Colocalization of GLUT4, Syt VII, and Kv1.3. Unstimulated adipocytes isolated from wild-type mice, maintained in primary culture, and examined by confocal microscopy. Colocalization of (A) GLUT4 and Syt VII, (B) Kv1.3 and GLUT4, (C) Kv1.3 and Syt VII.

Article Snippet: GLUT4 expression was detected in adipocytes by immunofluorescence (anti-GLUT4 antibody, Cell Signaling Technology, Beverly, MA).

Techniques: Isolation, Mouse Assay, Confocal Microscopy

Journal: PLoS ONE

Article Title: Dab1 (Disable Homolog-1) Reelin Adaptor Protein Is Overexpressed in the Olfactory Bulb at Early Postnatal Stages

doi: 10.1371/journal.pone.0026673

Figure Lengend Snippet: Relative levels of both Reln and Dab1 proteins by western blot in the olfactory bulb. (a) Reln and Dab1 protein bands identified in cytoplasm (CF) and nuclear (NF) fractions of wt and reeler mice. Identities of protein bands are indicated. Reln is absent in reeler . Dab1 quantities are different in CF and NF identified by distinct band intensities. (b) Amount of Dab1 compared between wt and reeler animals in both subcellular compartments. (c) Comparison of relative levels of Dab1 in CF and NF of wt and reeler mice. CF amount of Dab1 is statistically higher in wt at P15, while in reeler the levels are higher at P7 and P15. In NF, Dab1 levels are statistically higher exclusively at P15. (d) Comparison of Dab1 quantities between wt and reeler animals. Dab1 reeler content is significantly higher in CF at all studied ages. In NF this difference is restricted to P15. (* p

Article Snippet: Dab1 expression using two different anti-Dab1 antibodies by western blot and immunohistochemistry. (a) Western blot of wt OB extracts using two anti-Dab1 antibodies from Chemicon and Sigma-Aldrich.

Techniques: Western Blot, Mouse Assay

Journal: PLoS ONE

Article Title: Dab1 (Disable Homolog-1) Reelin Adaptor Protein Is Overexpressed in the Olfactory Bulb at Early Postnatal Stages

doi: 10.1371/journal.pone.0026673

Figure Lengend Snippet: Characterization of periglomerular Dab1 neurons with calbindin (CB), parvalbumin (PV) and tyrosine-hydroxylase (TH) markers. Green color corresponds to Dab1 expression, while the red one corresponds to either CB (a–b, g–h, m–n, s–t), PV (c–d, i––j, o–p, u–v) or TH (e–f, k–l, q–r, w–x) at P0 (a–f), P3 (g–l), P7 (m–r) and P15 (s–x). None CB (a–b) and PV (c–d) positive cells express Dab1 at P0–P3. By contrast, whole TH cell populations express nuclear Dab1 at these ages (e–f, g–i). At P7 some CB and PV are positive for Dab1. By contrary, in reeler mice PV cells do not express Dab1 and the cell number is lower (m–p). At this age, TH cells show a lack of Dab1 nuclear expression, which become apparent in some of TH-fibers (q–r). Similar results are observed at P15 (s–x). Nuclei counterstained with Hoechst (blue). GL, glomerular layer; EPL, external plexiform layer; MCL, mitral cell layer. Scale bar: 50 µm.

Article Snippet: Dab1 expression using two different anti-Dab1 antibodies by western blot and immunohistochemistry. (a) Western blot of wt OB extracts using two anti-Dab1 antibodies from Chemicon and Sigma-Aldrich.

Techniques: Expressing, Mouse Assay

Journal: PLoS ONE

Article Title: Dab1 (Disable Homolog-1) Reelin Adaptor Protein Is Overexpressed in the Olfactory Bulb at Early Postnatal Stages

doi: 10.1371/journal.pone.0026673

Figure Lengend Snippet: Identification of astroglial and oligodendroglial cell expressing Dab1. P0 (a–d), P3 (e–h), P7 (i–l) and P15 (m–p). Astrocytes are recognized by GFAP (a–b, e–f, i–j, m–n) whereas oligodendrocytes by RIP (c–d, g–h, k–l, o–p). At P0–P3, GFAP (red) is restricted to the ONL and GL (a–b, e–f) and onwards labels the remaining layers, being negative for Dab1 (i–j, m–n). RIP labeling (red) is restricted to non-myelin oligodendrocytes in the GcL at P0–P3 (c–d, g–h). At P7, RIP expression is located in myelin and non-myelin oligodendrocytes (k–l). Non-myelin oligodendrocytes are absent at P15 (o–p). In all cases oligodendrocytes are negatives for Dab1. Labeling pattern is indistinguishable between wt and reeler animals. Nuclei are counterstained with Hoechst (blue). ONL, olfactory nerve layer; GL, glomerular layer; EPL, external plexiform layer; MCL, mitral cells layer; GcL, granular cells layer. Scale bar: 100 µm.

Article Snippet: Dab1 expression using two different anti-Dab1 antibodies by western blot and immunohistochemistry. (a) Western blot of wt OB extracts using two anti-Dab1 antibodies from Chemicon and Sigma-Aldrich.

Techniques: Expressing, Labeling

Journal: PLoS ONE

Article Title: Dab1 (Disable Homolog-1) Reelin Adaptor Protein Is Overexpressed in the Olfactory Bulb at Early Postnatal Stages

doi: 10.1371/journal.pone.0026673

Figure Lengend Snippet: Colocalization of Dab1/Reln and Dab1/NeuN in postnatal olfactory bulb cells. Dab1 (green) and Reln (red) in wild type (a–d); Dab1 and NeuN (red) in wild type (e, h, k, n) and reeler (f, i, l, o). Nuclei are counterstained with Hoechst (blue). Images correspond to representative sagittal sections at P0 (a, e, f, g), P3, (b, h, i, j), P7 (c, k, l, m) and P15 (d, n, o, p). Reln shows a complementary location pattern with Dab1 positive nuclei of mitral cells (a–d). At P0, glomeruli are slightly apparent with Reln staining (a). From P3 to P15 glomeruli strongly express Dab1 and periglomerular cells start to express both Reln and nuclear Dab1 (b–d). From P3 the majority of granular and periglomerular cells are positive for NeuN (e–p), many of them with nuclear Dab1 expression (arrows). Cartoons represent the bilayer formation by the mitral cells (nuclear Dab1 staining) with respect to the superficial granular cells (NeuN labeling). This cell positioning is disrupted in reeler mice (cartoons). ONL, olfactory nerve layer; GL, glomerular layer; EPL, external plexiform layer; MCL, mitral cells layer; IPL, internal plexiform layer; GcL, granular cells layer; SEZ, subependymal zone. Scale bar: 100 µm.

Article Snippet: Dab1 expression using two different anti-Dab1 antibodies by western blot and immunohistochemistry. (a) Western blot of wt OB extracts using two anti-Dab1 antibodies from Chemicon and Sigma-Aldrich.

Techniques: Staining, Expressing, Labeling, Mouse Assay

Journal: PLoS ONE

Article Title: Dab1 (Disable Homolog-1) Reelin Adaptor Protein Is Overexpressed in the Olfactory Bulb at Early Postnatal Stages

doi: 10.1371/journal.pone.0026673

Figure Lengend Snippet: Pattern of Dab1 mRNA location in the postnatal olfactory bulb. Dab1 transcripts are shown in wt (a, c, e, g) and reeler (b, d, f, h) mice at P0 (a,b), P3, (c, d), P7 (e, f) and P15 (g, h). Transcripts are mainly detected in granular cells (black stars in GcL), mitral cells (white arrows in MCL) and several periglomerular cell populations and intra-glomeruli processes (black arrows in GL). No differences are observed between wt and reeler mice. Note the high presence of Dab1 mRNA in the rostral migratory stream (RMS) cells. Scale bar: 1 mm.

Article Snippet: Dab1 expression using two different anti-Dab1 antibodies by western blot and immunohistochemistry. (a) Western blot of wt OB extracts using two anti-Dab1 antibodies from Chemicon and Sigma-Aldrich.

Techniques: Mouse Assay

Journal: PLoS ONE

Article Title: Dab1 (Disable Homolog-1) Reelin Adaptor Protein Is Overexpressed in the Olfactory Bulb at Early Postnatal Stages

doi: 10.1371/journal.pone.0026673

Figure Lengend Snippet: Colocalization of Dab1/Map2a,b and Dab1/RC2 in postnatal olfactory bulb cells. Dab1 (green), Map2a,b (red) (a–b, f–g, k–l, p–q) and the radial glial cell marker RC2 (red) (c–e, h–j, m–o, r–t). OB sagittal sections at P0 (a–e), P3 (f–j), P7 (k–o) and P15 (p–t). At P0 neuronal Dab1 positive processes (Map2a,b) are evident in the GL, EPL and MCL (a, b) while radial glial Dab1 positive processes (RC2) occupied the OB surface (c–e). From P3 onwards, neuronal Dab1 positive neuronal fibers become evident in the IPL and GcL (f–g) while the radial cell processes decreased in quantity. Some Rc2 positive/Dab1 negative cells surround the glomeruli (h–j). Neuronal fibers pattern is similar at P7 (k–i) and P15 (p–q), whereas Rc2 fibers are absent from P7 (m–o). From P7 to P15 all radial glial cells are Dab1 negative (m–o; r–t). Nuclei counterstained with Hoechst (blue). ONL, olfactory nerve layer; GL, glomerular layer; EPL, external plexiform layer; MCL, mitral cells layer; IPL, internal plexiform layer; GcL, granular cells layer; SEZ, subependymal zone. Scale bar: 100 µm.

Article Snippet: Dab1 expression using two different anti-Dab1 antibodies by western blot and immunohistochemistry. (a) Western blot of wt OB extracts using two anti-Dab1 antibodies from Chemicon and Sigma-Aldrich.

Techniques: Marker

Journal: PLoS ONE

Article Title: Dab1 (Disable Homolog-1) Reelin Adaptor Protein Is Overexpressed in the Olfactory Bulb at Early Postnatal Stages

doi: 10.1371/journal.pone.0026673

Figure Lengend Snippet: Detection of Dab1 protein within cell nuclei. Labeling of isolated nuclei and tissue sections of the OB in wild type (a, c, e, g, i, k, m, o) and reeler (b, d, f, h, j, l, n, p) at P0 (a, b, e, f), P3 (c, d, g, h), P7 (i, j, m, n) and P15 (k, l, o, p). Labeling shows a green dotted pattern in nuclei from both isolated (a–d, i–l) and in vivo sections (e–h, m–p). Nuclei are counterstained with Hoechst (blue). In vivo images are single confocal sections from MCL (mitral cells layer). Scales bars: 10 µm in e–h, m–p; 5 µm in a–d, i–l.

Article Snippet: Dab1 expression using two different anti-Dab1 antibodies by western blot and immunohistochemistry. (a) Western blot of wt OB extracts using two anti-Dab1 antibodies from Chemicon and Sigma-Aldrich.

Techniques: Labeling, Isolation, In Vivo

Journal: PLoS ONE

Article Title: Dab1 (Disable Homolog-1) Reelin Adaptor Protein Is Overexpressed in the Olfactory Bulb at Early Postnatal Stages

doi: 10.1371/journal.pone.0026673

Figure Lengend Snippet: Dab1 protein expression in the postnatal olfactory bulb. Protein expression in wild type (a, c, e, g) and reeler (b, d, f, h) mice at P0 (a, b), P3, (c, d), P7 (e, f) and P15 (g, h) . Dab1 immunostaining is strongly localized in cell processes of the GL, EPL, and IPL. A dotted pattern is evident in GcL. Mitral and some periglomerular cells are easily identified by a characteristic nuclear Dab1 labeling. Nuclei labeling in periglomerular cells is evident from P3. No marked differences are observed between wt and reeler at these ages. Nuclei are counterstained with Hoechst (blue). Images represent sagittal sections of OB. Scale bars a-h: 100 µm; inset images: 1 mm.

Article Snippet: Dab1 expression using two different anti-Dab1 antibodies by western blot and immunohistochemistry. (a) Western blot of wt OB extracts using two anti-Dab1 antibodies from Chemicon and Sigma-Aldrich.

Techniques: Expressing, Mouse Assay, Immunostaining, Labeling