Journal: Histochemistry and Cell Biology

Article Title: Multispectral imaging reveals the tissue distribution of tetraspanins in human lymphoid organs

doi: 10.1007/s00418-015-1326-2

Figure Lengend Snippet: Expression of CD37 and CD53 on immune cell subsets in blood. a Flow cytometry analysis of expression of CD37 or CD53 ( black line ) on CD4 and CD8 T cells, B cells, monocytes and NK cells versus isotype control ( gray line ). Gating strategy is presented in Supplementary Figure 1. Expression levels of CD37 ( b ) and CD53 ( c ) were normalized for isotype staining by background subtraction. Experiments were performed with PBLs from three healthy donors. Data present mean ± SD. d Flow cytometry analysis of expression of CD37 or CD53 ( black line ) on mDCs (BDCA1 + CD19 − ) or pDCs (BDCA2 + ) versus isotype control ( gray line ). Expression levels of CD37 ( e ) and CD53 ( f ) were normalized for isotype staining by background subtraction. Experiments were performed with PBLs from two healthy donors. Data present mean ± SD

Article Snippet: Single-cell suspensions were first stained with primary antibodies against human CD53 (mem53, Serotec), CD37 (WR17, home-made) or isotype controls in PBS with 1 % BSA and 0.05 % NaN 3 (PBA) supplemented with 2 % HS for 30 min at 4 °C, followed by incubation with goat-anti-mouse Alexa488 antibody (Molecular Probes).

Techniques: Expressing, Flow Cytometry, Control, Staining

Journal: Histochemistry and Cell Biology

Article Title: Multispectral imaging reveals the tissue distribution of tetraspanins in human lymphoid organs

doi: 10.1007/s00418-015-1326-2

Figure Lengend Snippet: Subcellular localization of CD37 and CD53. Localization of a CD37 or b CD53 ( green ) in monocytes was studied by dual staining with calreticulin (ER), syntaxin 13 (endosomes) or Lamp1 (lysosomes) ( red ). Merge: co-localization in yellow ( white arrows ). Scale bar 5 μm

Article Snippet: Single-cell suspensions were first stained with primary antibodies against human CD53 (mem53, Serotec), CD37 (WR17, home-made) or isotype controls in PBS with 1 % BSA and 0.05 % NaN 3 (PBA) supplemented with 2 % HS for 30 min at 4 °C, followed by incubation with goat-anti-mouse Alexa488 antibody (Molecular Probes).

Techniques: Staining

Journal: Histochemistry and Cell Biology

Article Title: Multispectral imaging reveals the tissue distribution of tetraspanins in human lymphoid organs

doi: 10.1007/s00418-015-1326-2

Figure Lengend Snippet: Expression of CD37 and CD53 on human spleen. a Original multispectral image of human spleen stained for CD37 (Alexa488), CD53 (Alexa568) and cell nuclei (DAPI). Scale bar 100 μm. Composite RGB image ( b ) of unmixed CD37 ( c in green ), CD53 ( d in red ) and DAPI ( e in blue ) signal after correction for autofluorescence. One representative image is shown

Article Snippet: Single-cell suspensions were first stained with primary antibodies against human CD53 (mem53, Serotec), CD37 (WR17, home-made) or isotype controls in PBS with 1 % BSA and 0.05 % NaN 3 (PBA) supplemented with 2 % HS for 30 min at 4 °C, followed by incubation with goat-anti-mouse Alexa488 antibody (Molecular Probes).

Techniques: Expressing, Staining

Journal: Histochemistry and Cell Biology

Article Title: Multispectral imaging reveals the tissue distribution of tetraspanins in human lymphoid organs

doi: 10.1007/s00418-015-1326-2

Figure Lengend Snippet: Spectral imaging analysis of human spleen stained for CD20 (Warp Red), CD53 (True Blue) and cell nuclei (Nuclear Red). a A spectral library of three chromogens (Warp Red ( red line ), True Blue ( blue line ) and Nuclear Red ( green line )) was built in Nuance software using single-stained human spleen tissues. b Representative original multispectral image. Scale bar 100 μm. Composite RGB image ( c ) of unmixed CD20 ( d in red ), CD53 ( e in blue ) and nuclei ( f in green in composite RGB image) signal. g Tissue segmentation; B cell follicle (B, yellow ), red pulp (RP, green ) and other tissue (blood vessels, collagen; blue ). h Segmentation of individual cells ( green ) based on Nuclear Red staining. i Thresholds for Warp Red and True Blue staining were set to score CD20 − CD53 dim ( blue ), CD20 + CD53 dim ( red ), CD20 + CD53 bright ( yellow ) or CD20 − CD53 bright ( green ) cells. j–l Scatter plots showing optical densities for CD20 (Y-axis) and CD53 (X-axis) of individual cells in B cell follicles ( j , l blue ) and red pulp ( k–l red ) and thresholds used for scoring ( dotted lines ). A representative of 2000 cells per tissue region is plotted

Article Snippet: Single-cell suspensions were first stained with primary antibodies against human CD53 (mem53, Serotec), CD37 (WR17, home-made) or isotype controls in PBS with 1 % BSA and 0.05 % NaN 3 (PBA) supplemented with 2 % HS for 30 min at 4 °C, followed by incubation with goat-anti-mouse Alexa488 antibody (Molecular Probes).

Techniques: Imaging, Staining, Software

Journal: Histochemistry and Cell Biology

Article Title: Multispectral imaging reveals the tissue distribution of tetraspanins in human lymphoid organs

doi: 10.1007/s00418-015-1326-2

Figure Lengend Snippet: Localization and expression of CD53 in bone marrow ( a–f ), spleen ( g–l ) and appendix ( m–r ). a , g , m Representative original multispectral image of lymphoid organ stained for CD20 (Warp Red), CD53 (True Blue) and cell nuclei (Nuclear Red). Scale bars in a , g , m = 100 μm. b , h , n , Composite RGB image after spectral unmixing of original image ( red CD20, blue CD53, green nuclei). c , i , o , Image showing scoring of CD20 − CD53 dim ( blue ), CD20 + CD53 dim ( red ), CD20 + CD53 bright ( yellow ) or CD20 − CD53 bright ( green ) cells. d , j , p , Optical density of CD53 on individual cells in human bone marrow ( d ), in B cell follicle ( blue line ) and in T cell zone and in red pulp ( red line ) in human spleen ( j ) and in B cell follicle ( blue line ) and in lamina propria ( red line ) in human appendix ( p ). Optical densities were binned per 0.05 and normalized to % of max. Percentage of CD53 dim and CD53 bright cells in the CD20 + cell population ( e ) and in the CD3 + cell population ( f ) in human bone marrow. Percentage of CD53 bright cells in the CD20 + cell population in the B cell follicle and red pulp ( k ), and in the CD3 + cell population in the T cell zone and red pulp ( l ) in human spleen. Percentage of CD53 bright cells in the CD20+ cell population ( q ) and in the CD3 + cell population ( r ) in the B cell follicle and lamina propria in human appendix. Each dot represents data of one ×20 image from the lymphoid tissue. The red line represents the mean. * P < 0.05, **** P < 0.0001

Article Snippet: Single-cell suspensions were first stained with primary antibodies against human CD53 (mem53, Serotec), CD37 (WR17, home-made) or isotype controls in PBS with 1 % BSA and 0.05 % NaN 3 (PBA) supplemented with 2 % HS for 30 min at 4 °C, followed by incubation with goat-anti-mouse Alexa488 antibody (Molecular Probes).

Techniques: Expressing, Staining

Journal: Stem Cell Research & Therapy

Article Title: NCAPH-YAP1 interaction promotes breast cancer stemness and tumor progression

doi: 10.1186/s13287-025-04648-0

Figure Lengend Snippet: Transcriptome sequencing analysis of BC cell lines after NCAPH overexpression. A Heatmap of DEGs following NCAPH overexpression. B GSEA analysis showing enrichment of the Hippo signaling pathway. C KEGG pathway analysis of DEGs. D KMplot database analysis of the impact of YAP1 expression levels on OS in BC patients

Article Snippet: The primary antibodies used were against NCAPH (1:5000, Cat# 67,655–1-Ig), CD133 (1:5000, Cat#18,470–1-AP), CD44 (1:5000, Cat#60,224–1-Ig), YAP1 (1:5000, Cat# 13,584–1-AP), phospho-YAP1 (Ser127, 1:5000, Cat#29,018–1-AP), OCT4 (1:5000, Cat#60,242–1-Ig), SOX2 (1:2000, Cat#66,411–1-Ig), Nanog (1:2000, Cat#67,255–1-Ig), Cyclin D1 (1:5000, Cat#26,939–1-AP), CDK1 (1:5000, Cat#67,575–1-Ig), CDK2 (1:5000, Cat#10,122–1-AP), CDK4 (1:5000, Cat#66,950–1-Ig), β-actin (1:5000, Cat#66,009–1-Ig), Flag (1:5000, Cat#66,008–4-Ig), α-Tubulin (1:5000, Cat#66,031–1-Ig), and GAPDH (1:5000, Cat#10,494–1-AP), all sourced from Proteintech (Rosemont, IL, USA).

Techniques: Sequencing, Over Expression, Expressing

Journal: Stem Cell Research & Therapy

Article Title: NCAPH-YAP1 interaction promotes breast cancer stemness and tumor progression

doi: 10.1186/s13287-025-04648-0

Figure Lengend Snippet: NCAPH interacts with YAP1 and promotes YAP1 expression and nuclear translocation. A WB analysis examining the effects of NCAPH overexpression or knockdown on Hippo-YAP1 signaling (Full-length gels are presented in Supplementary Fig. 6). B WB analysis assessing the effect of NCAPH overexpression on YAP1 nuclear translocation (Full-length gels are presented in Supplementary Fig. 7). C IF showing co-localization of NCAPH and YAP1 proteins within cells. D Co-IP assay demonstrating the interaction between NCAPH and YAP1 (Full-length gels are presented in Supplementary Fig. 7)

Article Snippet: The primary antibodies used were against NCAPH (1:5000, Cat# 67,655–1-Ig), CD133 (1:5000, Cat#18,470–1-AP), CD44 (1:5000, Cat#60,224–1-Ig), YAP1 (1:5000, Cat# 13,584–1-AP), phospho-YAP1 (Ser127, 1:5000, Cat#29,018–1-AP), OCT4 (1:5000, Cat#60,242–1-Ig), SOX2 (1:2000, Cat#66,411–1-Ig), Nanog (1:2000, Cat#67,255–1-Ig), Cyclin D1 (1:5000, Cat#26,939–1-AP), CDK1 (1:5000, Cat#67,575–1-Ig), CDK2 (1:5000, Cat#10,122–1-AP), CDK4 (1:5000, Cat#66,950–1-Ig), β-actin (1:5000, Cat#66,009–1-Ig), Flag (1:5000, Cat#66,008–4-Ig), α-Tubulin (1:5000, Cat#66,031–1-Ig), and GAPDH (1:5000, Cat#10,494–1-AP), all sourced from Proteintech (Rosemont, IL, USA).

Techniques: Expressing, Translocation Assay, Over Expression, Knockdown, Co-Immunoprecipitation Assay

Journal: Stem Cell Research & Therapy

Article Title: NCAPH-YAP1 interaction promotes breast cancer stemness and tumor progression

doi: 10.1186/s13287-025-04648-0

Figure Lengend Snippet: The YAP1 inhibitor Verteporfin reverses NCAPH overexpression-induced enhancement of CSC properties, proliferation, migration, and invasion in BC cells. A Verteporfin inhibits YAP1 expression and also reverses the increase in tumorsphere formation induced by NCAPH overexpression (Full-length gels are presented in Supplementary Fig. 8). B WB experiments show that Verteporfin reverses CSC marker protein expression induced by NCAPH overexpression (Full-length gels are presented in Supplementary Fig. 8). C The CCK-8 assay shows that Verteporfin reverses the enhanced proliferative capacity in BC cell lines induced by NCAPH overexpression. D ​The wound healing assay indicates that Verteporfin reverses the increase in migratory capacity of BC cell lines induced by NCAPH overexpression. E The Transwell assay reveals that Verteporfin markedly reverses the enhancement of migratory and invasive capabilities in BC cell lines induced by NCAPH overexpression. Data are presented as mean ± SD (*P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001)

Article Snippet: The primary antibodies used were against NCAPH (1:5000, Cat# 67,655–1-Ig), CD133 (1:5000, Cat#18,470–1-AP), CD44 (1:5000, Cat#60,224–1-Ig), YAP1 (1:5000, Cat# 13,584–1-AP), phospho-YAP1 (Ser127, 1:5000, Cat#29,018–1-AP), OCT4 (1:5000, Cat#60,242–1-Ig), SOX2 (1:2000, Cat#66,411–1-Ig), Nanog (1:2000, Cat#67,255–1-Ig), Cyclin D1 (1:5000, Cat#26,939–1-AP), CDK1 (1:5000, Cat#67,575–1-Ig), CDK2 (1:5000, Cat#10,122–1-AP), CDK4 (1:5000, Cat#66,950–1-Ig), β-actin (1:5000, Cat#66,009–1-Ig), Flag (1:5000, Cat#66,008–4-Ig), α-Tubulin (1:5000, Cat#66,031–1-Ig), and GAPDH (1:5000, Cat#10,494–1-AP), all sourced from Proteintech (Rosemont, IL, USA).

Techniques: Over Expression, Migration, Expressing, Marker, CCK-8 Assay, Wound Healing Assay, Transwell Assay

Journal: Military Medical Research

Article Title: Endophilin A2 controls touch and mechanical allodynia via kinesin-mediated Piezo2 trafficking.

doi: 10.1186/s40779-024-00520-z

Figure Lengend Snippet: Fig. 9 Hypothetical model illustrating that endophilin A2 (EndoA2) interacts with Piezo2 and KIF5B to form a complex to regulate the membrane trafficking of Piezo2 in sensory neurons and thus contributes to mechanical hypersensitivity

Article Snippet: DRG neurons from L4–L6 of mice were cultured for 2 d. Following fixation, DRG coverslips were treated with primary antibodies against Piezo2 (rabbit, 1:200, Novus, USA, NBP1-78,624) and NF200 (mouse, 1:200, Sigma, USA, N0142) and the appropriate secondary antibodies.

Techniques: Membrane

Journal: Cancers

Article Title: Proteome-Wide Analysis Reveals TFEB Targets for Establishment of a Prognostic Signature to Predict Clinical Outcomes of Colorectal Cancer.

doi: 10.3390/cancers15030744

Figure Lengend Snippet: Figure 1. Mutations in S138 and S142 caused almost complete nuclear retention of TFEB. (A) Schematic of the human TFEB protein domains. S138 and S142 are localized in the proxim- ity of a NES. (B) S138 and S142 of TFEB are evolutionarily conserved in the indicated species. Alignment of the sequences near TFEB S138 and S142 is shown. (C) Western blots of endogenous and exogenous TFEB in CRC cells transfected with wild-type TFEB-GFP or TFEB-GFP mutant (S142A/S138A). Uncropped immunoblots are provided in the Figure S1. (D,E) Localization of TFEBWT and TFEBS142A/S138A in the cytosol and in the nucleus was detected by confocal microscopy. Representative images (D) and statistical results (E) are shown. Torin1 (250 nm), a positive control. Cell outlines in white dotted lines. DAPI was used to label the nucleus. Scale bars, 5 µm. Mean ± SD, n = 10 cells per condition, unpaired t-test. (F) Cell viability of indicated cells were analyzed using a CCK8 assay. Mean ± SEM, n = 3, unpaired t-test. ** p < 0.01, *** p < 0.001.

Article Snippet: The primary antibodies (against TFEB, GFP, LAMP2, NDRG1, SMPD1, and actin) and the HRP-conjugated secondary antibodies (goat anti-rabbit antibody) were supplied by Proteintech (Wuhan, China).

Techniques: Western Blot, Transfection, Mutagenesis, Confocal Microscopy, Positive Control, CCK-8 Assay

Journal: International Journal of Molecular Sciences

Article Title: Palmitoylethanolamide Inhibits Glutamate Release in Rat Cerebrocortical Nerve Terminals

doi: 10.3390/ijms16035555

Figure Lengend Snippet: PEA-mediated inhibition of evoked glutamate release is partially blocked by the selective cannabinoid CB1 receptor antagonist AM281. Synaptosomes were fixed onto polylysine-coated coverslips and double stained for immunocytochemistry with antisera against cannabinoid CB1 receptors (red in ( a )) and the vesicular marker synaptophysin (green in ( b )). Merged panels are shown in ( c ) (orange). Scale bar, 10 μm; ( d ) Bar graph showing glutamate release induced by 1 mM 4-aminopyridine in the absence (control) or presence of 5 μM PEA, 10 μM AM281 (an antagonist of cannabinoid CB1 receptor), 10 μM AM281 and 5 μM PEA; 5 μM WIN55212-2 (an agonist of cannabinoid CB1 receptor), 5 μM WIN55212-2 and 5 μM PEA; 10 μM capsazepine (an antagonist of TRPV1); 10 μM capsazepine and 5 μM PEA; 10 μM GW6471 (an antagonist of PPARα); or 10 μM GW6471 and 5 μM PEA. Results are mean ± SEM of 5–7 independent experiments. *** p < 0.001 versus control group. # p < 0.05 versus the AM281-treated group.

Article Snippet: They were then incubated with a mixture of primary mouse monoclonal antibodies against synaptophysin (1:200; Abcam, Cambridge, UK) and rabbit monoclonal antibodies against cannabinoid CB 1 receptor (1:100; Abcam) for 90 min at room temperature.

Techniques: Inhibition, Staining, Immunocytochemistry, Marker