16-9381-81

antibody from Invitrogen Antibodies

Targeting: PDPN aggrus, Gp38, GP40, PA2.26, T1A-2

Provider product page for 16-9381-81

Flow cytometry

Other assay

Antibody data

Antibody Data
Antigen structure
References [7]
Comments [0]
Validations
Flow cytometry [1]
Other assay [10]

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Product number: 16-9381-81 - Provider product page
Provider: Invitrogen Antibodies
Product name: Podoplanin Monoclonal Antibody (NZ-1.3), Functional Grade, eBioscience™
Antibody type: Monoclonal
Antigen: Other
Description: Description: Podoplanin is a 40-kDa type I transmembrane sialomucin-like glycoprotein involved in platelet aggregation and cancer metastasis. This protein is expressed on lymphatic endothelial cells and follicular dendritic cells, as well as on many human cancers, including squamous cell carcinoma, glioblastoma, seminoma, and mesothelioma. Podoplanin expression has been demonstrated to be regulated by the Fos protein of the Activating Protein-1 (AP-1) transcription factor complex. Much of its functionality arises via interaction with its endogenous receptor, C-type lectin-like receptor-2 (CLEC-2). This antibody has been reported to block the interaction between Podoplanin and CLEC-2. Applications Reported: This NZ-1.3 antibody has been reported for use in flow cytometric analysis. Applications Tested: This NZ-1.3 antibody has been tested by flow cytometric analysis of the U87 cell line. This can be used at less than or equal to 0.5 µg per test. A test is defined as the amount (µg) of antibody that will stain a cell sample in a final volume of 100 µL. Cell number should be determined empirically but can range from 10^5 to 10^8 cells/test. It is recommended that the antibody be carefully titrated for optimal performance in the assay of interest. Storage and handling: Use in a sterile environment. Filtration: 0.2 µm post-manufacturing filtered. Purity: Greater than 90%, as determined by SDS-PAGE. Endotoxin Level: Less than 0.001 ng/µg antibody, as determined by LAL assay. Aggregation: Less than 10%, as determined by HPLC.
Reactivity: Human
Host: Rat
Isotype: IgG
Antibody clone number: NZ-1.3
Vial size: 50 µg
Concentration: 1 mg/mL
Storage: 4° C

PDPN protein structure - 16-9381-81 shown in red.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Fig 4 Human extravillous trophoblast cells invade decidual veins and lymphatics early during first trimester. (A) Histological sections of decidua basalis from a (mosaic) Klinefelter syndrome (47,XXY and 46, XY) pregnancy at W8.4. Left panels show low magnifications of same section immunostained for pKRT and PECAM1 (top) and chrX/chrY FISH (bottom). Middle-left and middle-right panels are magnifications of the (numbered) dashed boxes in the left panels. Right panels are magnifications of the dashed boxes in the middle-right panels. White arrowheads point to XXY fetal EVTs. (B) Histological sections of decidua basalis at W7.2, W8.4 and W10 immunostained for pKRT, PDPN and PECAM1 (left panels). FISH for chrX and chrY (right) was performed in the in the pKRT/PDPN/PECAM1-stained sections. White arrows point to EVTs invading the lymphatic vessels. All scale bars are depicted.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Fig 5 EVT aggregates and syncytial knots in the decidua basalis and parietalis. (A) Histological sections of decidua basalis at W5.5 and W7.5 immunostained for pKRT, PDPN and PECAM1 (lefts panels) showing the presence of mononuclear EVT aggregates (top row) and syncytial knots (bottom row). FISH for chrX and chrY (right panels) was performed in pKRT/PDPN/PECAM1-stained sections. White arrow indicates PDPN-positive lymphatics. (B) Histological sections of W10.6 decidua basalis (top row) and W8.1 decidua parietalis (bottom row) immunostained for pKRT and PECAM1 showing the presence of syncytial knots in veins. FISH for chrX and chrY (right panels) was performed in pKRT/PECAM1-stained sections. All scale bars are depicted.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: 1 FIGURE Identification of iAT1 cells in FD-AOs. A, A schematic representation of the analysis of scRNA-seq in FD-AOs. FD-EpCAM + cells (P2) were collected from two independent FD-AOs using FACS, followed by single-cell isolation using a Fluidigm C1 system and RNA sequencing. Bioinformatics data were processed using Seurat (version 3.1.2) including FD-SFTPC + cells (P2) (GSE90813). B, Flow cytometric analyses of FD-AOs (P2) with or without treatment with DMSO from day 2 to 14 before being subjected to scRNA-seq. C, UMAP plots displaying cell clusters of FD-EpCAM + cells (P2) in FD-AOs. We defined five major types of FD-EpCAM + cells: iAT1 cells, FD-iAT2 cells, mitotic cells, ASCL1 + PNECs, and NKX2-1 + cells. D, UMAP plots showing the expression of the selected marker genes; AT1 cells ( AGER , CAV1 , and PDPN ), AT2 cells ( SFTPB and SFTPC ), mitotic cells ( TOP2A ), ASCL1 + PNECs ( ASCL1 ), NKX2-1 + cells ( NKX2-1 ), and alveolar epithelial cells ( CPM ). E, Violin plots showing the gene expression distributions of the selected marker genes across cell types. F,G, Immunostaining of AT1 markers (HT1-56, PDPN, and AGER) and SFTPC-GFP in FD-AOs (P3). Scale bars = 10 mum. DMSO, dimethyl sulfoxide; FACS, fluorescence-activated cell sorting; FD-AOs, fibroblast-dependent alveolar organoids; scRNA-seq, single-cell RNA sequencing; UMAP, uniform manifold approximation and projection

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: 2 FIGURE iAT1 cells are present in hiPSC-derived EpCAM + cell population in FD-AOs but poorly detected in FF-AOs. A, A schematic representation of the analysis of scRNA-seq in FD-AOs and FF-AOs. FD-EpCAM + cells (P0) and FF-EpCAM + cells (P0) were isolated using FACS. Next, scRNA-seq was performed and sequencing data, including that of FD-EpCAM + cells (P2), FD-EpCAM + /DMSO cells (P2), and FD-SFTPC + cells (P2), were analyzed. B, Flow cytometric analyses of FD-EPCAM + cells (P0) and FF-EpCAM + cells (P0) used for scRNA-seq. C, UMAP plots displaying cell clusters in FD-AOs and FF-AOs. We defined eight major types of hiPSC-derived EpCAM + cells: iAT1 cells, FD-iAT2 cells, mitotic cells, ASCL1 + PNECs, ID1 + cells, NKX2-1 + cells, gastric-like cells, and FF-EpCAM + cells. D, Violin plots showing the gene expression distributions of each representative marker gene. E, Immunostaining of AT1 markers (HT1-56, PDPN, and/or AGER) and SFTPC-GFP in the FD-AO (P3) and FF-AO (P0). Scale bars = 10 mum. DMSO, dimethyl sulfoxide; FACS, fluorescence-activated cell sorting; FD-AOs, fibroblast-dependent alveolar organoids; FF-AOs, fibroblast-free alveolar organoids; hiPSC, human induced pluripotent stem cell; scRNA-seq, single-cell RNA sequencing; UMAP, uniform manifold approximation and projection

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: 6 FIGURE XAV-939 increases the population of iAT1 cells in FD-AOs. A, A schematic diagram of the analysis of the effect of XAV-939 on FD-AOs. B,C, Flow cytometric analyses of the PDPN and SFTPC expression of FD-EpCAM + cells (P2-5). FD-AOs (P2-5) were treated with 10 muM XAV-939 or DMSO from day 2 to 14, and then the proportion of SFPTC - PDPN + cells in FD-AOs was measured on day 14 (mean +- SEM, n = 3 from three independent experiments). The values indicate the percentage of each population. * P < .05 vs DMSO-treated hiPSC-derived SFTPC - PDPN + cells (unpaired two-tailed Student's t test). D,E, Representative bright-field and SFTPC-GFP fluorescence microscopic live cell images of FD-AOs (P2-5) treated with DMSO or 10 muM XAV-939 from day 2 to 14. Thickness of each FD-AO (P2-P5) treated with DMSO or 10 muM XAV-939 was measured (mean +- SEM, n = 18 spheres from three independent experiments). Scale bars = 50 mum. * P < .05 vs DMSO-treated FD-AO (unpaired two-tailed Student's t test). F, Immunostaining of AT1 markers (AGER, HOPX, HT1-56, and/or PDPN) and SFTPC-GFP in DMSO- or XAV-939-treated FD-AOs (P3). Scale bars = 10 mum. DMSO, dimethyl sulfoxide; FD-AOs, fibroblast-dependent alveolar organoids; hiPSC, human induced pluripotent stem cell

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Fig. 4 Identification of CADM1 as a phenotypic marker of eSSPCs. a Dot plots showing the expression of differentially expressed cell surface genes (left) and candidate SSC markers (right) in 8 WPC human long bone subsets. Asterisks indicated positive markers that were used to enrich eSSPCs. b Immunofluorescent images of PDPN + CADM1 + cells in 8 WPC human long bones. Overviews of PDPN + CADM1 + cells (arrows) in the articular (upper left) and POC (bottom left) regions were shown on the left. PDPN + CADM1 + cells were found in the inner layer of perichondrium in the articular regions (I) and surrounding POC (II). A few PDPN + CADM1 + cells were also found inside POC (III). Arrow heads indicated enlarged PDPN + CADM1 + cells. Merged and single-channel images of DAPI (blue), CADM1 (red) and PDPN (green) were shown ( n = 2 embryos). Scale bars in snapshot images, 50 mum; scale bars in magnified images, 5 mum. c Flow cytometry gating strategies for sorting different populations in 8 WPC long bones ( n = 3 embryos). d Representative crystal violet staining of CFU-F colonies generated by the sorted populations as indicated in c . Magnified images of the boxed areas were shown on the right. Scale bars, 25 mum. e Quantifications of the number (left) and mean diameter (right) of the CFU-F colonies ( n = 3 embryos). The statistical significance of differences was determined using one-way ANOVA with multiple comparison tests (LSD). * P < 0.05; ** P < 0.01; *** P < 0.001. Error bars indic

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Fig. 5 Functional characterizations of eSSPCs in vitro and in vivo. a Flow cytometry plots showing the maintenance of phenotypic eSSPCs after serially passaging clonally expanded PDGFRA low/- PDPN + CADM1 + cells ( n = 3 clones). b Representative oil red O (top), alizarin red (middle) and toluidine blue (bottom) staining after adipogenic, osteogenic and chondrogenic differentiation of clonally expanded eSSPCs (PDGFRA low/- PDPN + CADM1 + ). Magnified images of the boxed areas were shown on the right. Scale bars, 200 mum. c qPCR analyses of adipogenic, osteogenic and chondrogenic marker genes in clonally expanded eSSPCs before and after trilineage differentiation in vitro ( n = 3 clones). The statistical significance of differences was determined using Wilcoxon signed rank test. * P < 0.05; ** P < 0.01. Error bars indicated SEM. d Renal subcapsular transplantation. The work flow for functional characterization of eSSPC in vivo (top). Subcapsular xenografts were dissected and sectioned 8 weeks after transplantation of culture expanded eSSPCs into immunodeficient mice. Bright field (middle), Movat pentachrome staining (bottom left; cartilage, blue; bone and fibrous tissue, yellow) and immunofluorescent staining images (bottom right; DAPI, blue; collagen I (COL1), red; collagen II (COL2), green) were shown ( n = 9 grafts from three embryos). Scale bars, 50 mum.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Figure 2 Characterization of ECs by flow cytometry markers. The cells were stained for the following markers: VWF, ACE, CD31, CD34, CD133, CD105, PDPN, AP, alphaSMA, EGFR. Data were recorded for 10,000 events using CellQuest software (v.2.3.0.84) and presented as histogram overlays. ( A ) The most characteristic markers for ECs and EPCs. ( B ) Markers associated with cancerous phenotype. Histogram overlays display representative repetitions; gray--healthy ECs unstained, black--tumor ECs unstained, green--healthy ECs stained, red--tumor ECs stained. Y axis = the number of events; X axis = fluorescence intensity; the bar charts present delta MFI. Ns--not significant; * p < 0.05 in Student t test vs. HBH.MEC. Data are reported as the means +- SEM (n = 3).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: 10.1371/journal.ppat.1009600.g005 Fig 5 Characterization of KSHV-infected PDLSCs of distinct states in the mesenchymal-to-endothelial differentiation spectrum. (A) Mock- and KSHV-infected PDLSCs and LECs were examined for PDGFRA and PDPN expression profile by flow cytometry analysis. Three subpopulations (xM, hybrid M/E, and xE) were quantified based on the PDGFRA and PDPN profiles ( n = 3 independent experiments). Statistical analyses were performed using two-tailed Student's test and P-values were calculated by GraphPad Prism. *p < 0.05, **p < 0.01, ***p < 0.001. ( B) PDGFRA and PDPN expression in PDLSC and K-PDLSC spheroids at 4 days post-infection were analyzed by IFA. Scale bar, 50 mum. ( C) K-PDLSCs were stained for PDGFRA and PDPN and sorted by flow cytometry. The purified xM and M/E populations were cultured for 10 days and their PDGFRA/PDPN profiles were examined for their phenotypic plasticity. ( D) Western blot analysis of xM, M/E, and LECs for their mesenchymal and endothelial markers. ( E) The expression profiles of mesenchymal and endothelial markers in hybrid M/E and xM state cells were analyzed at the mRNA level by RT-qPCR. ( F) IFA analysis of LANA, TAGLN, and VCAM in xM and M/E state cells. Scale bar, 50 mum.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Cell surface analysis of AMSCs within the transitional culture AMSCs were cultured in a 10% collagen type I gel for 7 days and embedded in a 0.2% collagen type I gel, making up the transitional culture for an additional 14 days. Flow cytometric analysis was conducted for PDPN, CD73, and CD146 (representative data presented).