anti aqp3  (Alomone Labs)


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    Structured Review

    Alomone Labs anti aqp3
    <t>AQP3</t> Immunoreactivity was Mislocalized in a Majority of Psoriatic Lesions (A through C) Archived paraffin-embedded psoriasis samples were sectioned (4 μm) and deparaffinized. Sections were then stained with an antibody recognizing AQP3 and visualized with an ABC kit and DAB. Results from three patients are shown and the staining pattern is representative of 8 of 10 psoriatic samples examined. (D) A negative control was performed by omitting primary antibody. Note that AQP3 is localized to the cytoplasm rather than to the plasma membrane in a majority of the psoriatic epidermis. All sections were counterstained with hematoxylin.
    Anti Aqp3, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 93/100, based on 9 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/anti aqp3/product/Alomone Labs
    Average 93 stars, based on 9 article reviews
    Price from $9.99 to $1999.99
    anti aqp3 - by Bioz Stars, 2022-10
    93/100 stars

    Images

    1) Product Images from "ABNORMAL AQUAPORIN-3 PROTEIN EXPRESSION IN HYPERPROLIFERATIVE SKIN DISORDERS"

    Article Title: ABNORMAL AQUAPORIN-3 PROTEIN EXPRESSION IN HYPERPROLIFERATIVE SKIN DISORDERS

    Journal: Archives of dermatological research

    doi: 10.1007/s00403-011-1136-x

    AQP3 Immunoreactivity was Mislocalized in a Majority of Psoriatic Lesions (A through C) Archived paraffin-embedded psoriasis samples were sectioned (4 μm) and deparaffinized. Sections were then stained with an antibody recognizing AQP3 and visualized with an ABC kit and DAB. Results from three patients are shown and the staining pattern is representative of 8 of 10 psoriatic samples examined. (D) A negative control was performed by omitting primary antibody. Note that AQP3 is localized to the cytoplasm rather than to the plasma membrane in a majority of the psoriatic epidermis. All sections were counterstained with hematoxylin.
    Figure Legend Snippet: AQP3 Immunoreactivity was Mislocalized in a Majority of Psoriatic Lesions (A through C) Archived paraffin-embedded psoriasis samples were sectioned (4 μm) and deparaffinized. Sections were then stained with an antibody recognizing AQP3 and visualized with an ABC kit and DAB. Results from three patients are shown and the staining pattern is representative of 8 of 10 psoriatic samples examined. (D) A negative control was performed by omitting primary antibody. Note that AQP3 is localized to the cytoplasm rather than to the plasma membrane in a majority of the psoriatic epidermis. All sections were counterstained with hematoxylin.

    Techniques Used: Staining, Negative Control

    AQP3 Immunoreactivity was “Patchy” in Squamous Cell Carcinoma (SCC) Archived paraffin-embedded squamous cell carcinoma (SCC) samples were sectioned (4 μm) and deparaffinized. Sections were then stained with antibodies recognizing AQP3 and visualized with an ABC kit and DAB. Sections were counterstained with hematoxylin. Illustrated in panels A through C is AQP3 staining in 3 patients with results representative of 5 of 5 SCCs. A negative control in which the primary antibody was omitted showed no AQP3 staining (panel D).
    Figure Legend Snippet: AQP3 Immunoreactivity was “Patchy” in Squamous Cell Carcinoma (SCC) Archived paraffin-embedded squamous cell carcinoma (SCC) samples were sectioned (4 μm) and deparaffinized. Sections were then stained with antibodies recognizing AQP3 and visualized with an ABC kit and DAB. Sections were counterstained with hematoxylin. Illustrated in panels A through C is AQP3 staining in 3 patients with results representative of 5 of 5 SCCs. A negative control in which the primary antibody was omitted showed no AQP3 staining (panel D).

    Techniques Used: Staining, Negative Control

    AQP3 Staining was Reduced in Ki67-Positive Cells in Squamous Cell Carcinoma Archived paraffin-embedded squamous cell carcinoma (SCC) samples were sectioned (4 μm) and deparaffinized. Sequential serial sections were then stained with antibodies recognizing (A, C) AQP3 or (B, D) Ki67 [an antigen expressed in proliferating cells (Dako, Carpinteria, CA)] and visualized with an ABC kit and DAB. Sections were counterstained with hematoxylin. Note that cells that are Ki67 positive exhibit reduced or absent AQP3 staining (arrows in panels C and D). The inverse correlation of Ki67 positivity with AQP3 immunoreactivity is representative of 4 of 4 SCCs. (E) Human tonsil stained with Ki67 as a positive control. (F) Primary antibody was omitted as a negative control.
    Figure Legend Snippet: AQP3 Staining was Reduced in Ki67-Positive Cells in Squamous Cell Carcinoma Archived paraffin-embedded squamous cell carcinoma (SCC) samples were sectioned (4 μm) and deparaffinized. Sequential serial sections were then stained with antibodies recognizing (A, C) AQP3 or (B, D) Ki67 [an antigen expressed in proliferating cells (Dako, Carpinteria, CA)] and visualized with an ABC kit and DAB. Sections were counterstained with hematoxylin. Note that cells that are Ki67 positive exhibit reduced or absent AQP3 staining (arrows in panels C and D). The inverse correlation of Ki67 positivity with AQP3 immunoreactivity is representative of 4 of 4 SCCs. (E) Human tonsil stained with Ki67 as a positive control. (F) Primary antibody was omitted as a negative control.

    Techniques Used: Staining, Positive Control, Negative Control

    AQP3 Immunoreactivity was Reduced or Absent in Basal Cell Carcinoma (BCC) Archived paraffin-embedded basal cell carcinoma samples were sectioned (4 μm), deparaffinized, stained with antibodies recognizing AQP3 and visualized with an ABC kit and 3,3’-diaminobenzidine (DAB). Sections were counterstained with hematoxylin. Results illustrate three BCCs from different patients and are representative of 13 of 13 BCCs. The results were quantified as 3+ (intense staining), 2+ (moderate staining), + (weak staining) or 0 (no staining) by three independent observers and the values averaged. Shown in panel D is the quantitation (means ± SEM) of the 13 BCCs relative to normal-appearing overlying epidermis with the data analyzed using a student's t-test; *p
    Figure Legend Snippet: AQP3 Immunoreactivity was Reduced or Absent in Basal Cell Carcinoma (BCC) Archived paraffin-embedded basal cell carcinoma samples were sectioned (4 μm), deparaffinized, stained with antibodies recognizing AQP3 and visualized with an ABC kit and 3,3’-diaminobenzidine (DAB). Sections were counterstained with hematoxylin. Results illustrate three BCCs from different patients and are representative of 13 of 13 BCCs. The results were quantified as 3+ (intense staining), 2+ (moderate staining), + (weak staining) or 0 (no staining) by three independent observers and the values averaged. Shown in panel D is the quantitation (means ± SEM) of the 13 BCCs relative to normal-appearing overlying epidermis with the data analyzed using a student's t-test; *p

    Techniques Used: Staining, Quantitation Assay

    2) Product Images from "Differential roles of VPS and RAAS in water homeostasis and a risk for kidney dysfunction in rats undergoing rapid fasting/dehydration with regular exercise"

    Article Title: Differential roles of VPS and RAAS in water homeostasis and a risk for kidney dysfunction in rats undergoing rapid fasting/dehydration with regular exercise

    Journal: Physiological Reports

    doi: 10.14814/phy2.14670

    Effects of the rapid restriction of food and water intake on the renal expression levels of water channels in rats with and without regular exercise. (a) Gene expression levels of AQP2 in the kidney medulla. The mRNA levels of AQP2 were obtained by quantitative RT‐PCR and normalized to those of GAPDH. Expression levels are displayed relative to those of the control group receiving neither regular exercise nor rapid restriction (1.0). (b) Protein levels of AQP2 in the kidney medulla. The amounts of the nonglycosylated (29 kDa) and glycosylated form (35 kDa) forms of AQP2 protein were analyzed by Western blot (left panel) and densitometry, and the sum of the two values was compared between the groups (right panel). The image shows representative immunoblots. (c–f) Gene expression levels and protein levels of AQP3 (c and d, respectively) and AQP4 (e and f, respectively) in the kidney medulla. Data are obtained and displayed as described in (a and b). Data are expressed as mean ± SEM (mRNA levels, n = 5/group; protein levels, n = 4/group). * p
    Figure Legend Snippet: Effects of the rapid restriction of food and water intake on the renal expression levels of water channels in rats with and without regular exercise. (a) Gene expression levels of AQP2 in the kidney medulla. The mRNA levels of AQP2 were obtained by quantitative RT‐PCR and normalized to those of GAPDH. Expression levels are displayed relative to those of the control group receiving neither regular exercise nor rapid restriction (1.0). (b) Protein levels of AQP2 in the kidney medulla. The amounts of the nonglycosylated (29 kDa) and glycosylated form (35 kDa) forms of AQP2 protein were analyzed by Western blot (left panel) and densitometry, and the sum of the two values was compared between the groups (right panel). The image shows representative immunoblots. (c–f) Gene expression levels and protein levels of AQP3 (c and d, respectively) and AQP4 (e and f, respectively) in the kidney medulla. Data are obtained and displayed as described in (a and b). Data are expressed as mean ± SEM (mRNA levels, n = 5/group; protein levels, n = 4/group). * p

    Techniques Used: Expressing, Quantitative RT-PCR, Western Blot

    3) Product Images from "Empagliflozin Contributes to Polyuria via Regulation of Sodium Transporters and Water Channels in Diabetic Rat Kidneys"

    Article Title: Empagliflozin Contributes to Polyuria via Regulation of Sodium Transporters and Water Channels in Diabetic Rat Kidneys

    Journal: Frontiers in Physiology

    doi: 10.3389/fphys.2019.00271

    Empagliflozin causes no significant changes to AQP3 and AQP1 but tends to increase AQP7 level in diabetic rat kidneys. (A) Representative renal sections immunostained with anti-AQP3. (B) Quantitative analysis of results for AQP3 shows no significant change among groups. Magnification, ×200. (C) Representative immunoblot reacting with anti-NCC, anti-α-ENaC and anti- γ-ENaC. (D) Densitometric analysis shows that the expression of AQP3 protein significantly increased in all OLETF rats. Lixisenatide treatment further increased the expression of AQP3. ∗ P
    Figure Legend Snippet: Empagliflozin causes no significant changes to AQP3 and AQP1 but tends to increase AQP7 level in diabetic rat kidneys. (A) Representative renal sections immunostained with anti-AQP3. (B) Quantitative analysis of results for AQP3 shows no significant change among groups. Magnification, ×200. (C) Representative immunoblot reacting with anti-NCC, anti-α-ENaC and anti- γ-ENaC. (D) Densitometric analysis shows that the expression of AQP3 protein significantly increased in all OLETF rats. Lixisenatide treatment further increased the expression of AQP3. ∗ P

    Techniques Used: Expressing

    4) Product Images from "Aquaporin-3 regulates endosome-to-cytosol transfer via lipid peroxidation for cross presentation"

    Article Title: Aquaporin-3 regulates endosome-to-cytosol transfer via lipid peroxidation for cross presentation

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0238484

    AQP3 regulates phagosomal lipid peroxidation. ( A ) AQP3 -/- or WT BMDCs were co-incubated with fluorescently-labeled OVA-coated 1 μm beads and a C11-bodipy lipid peroxidation indicator for 1.5 hours, homogenized, and phagosomes were analyzed by flow cytometry. ( B ) Annotated volcano plot of ECT functional genomics screen highlights the connection between ECT and ROS generation and transport, which could potentiate endosomal lipid peroxidation and antigen release. ( C ) ECT was evaluated in WT BMDCs with 0.5 mg/ml β-lactamase in media that did or did not contain ISCOMATRIX ™ adjuvant. ( D ) WT BMDCs were incubated with 100 mg/ml OVA in media that did or did not contain ISCOMATRIX ™ adjuvant, in the presence or absence of 1 μM epoxomicin, followed by co-incubation with CFSE-labeled OT-I CD8+ T cells for 64 hours. The number of CD8+ T cells that had undergone division as a measure of cross presentation was evaluated by flow cytometry. ( E ) DHFR activity of recombinant β-lactamase-DHFR fusion protein was measured after incubation with 500 nM MTX or equivalent volume PBS. ( F ) The ability of increasing amounts of recombinant β-lactamase-DHFR fusion protein to undergo ECT was evaluated in WT BMDCs following treatment with 500 nM MTX or equivalent volume PBS. *P
    Figure Legend Snippet: AQP3 regulates phagosomal lipid peroxidation. ( A ) AQP3 -/- or WT BMDCs were co-incubated with fluorescently-labeled OVA-coated 1 μm beads and a C11-bodipy lipid peroxidation indicator for 1.5 hours, homogenized, and phagosomes were analyzed by flow cytometry. ( B ) Annotated volcano plot of ECT functional genomics screen highlights the connection between ECT and ROS generation and transport, which could potentiate endosomal lipid peroxidation and antigen release. ( C ) ECT was evaluated in WT BMDCs with 0.5 mg/ml β-lactamase in media that did or did not contain ISCOMATRIX ™ adjuvant. ( D ) WT BMDCs were incubated with 100 mg/ml OVA in media that did or did not contain ISCOMATRIX ™ adjuvant, in the presence or absence of 1 μM epoxomicin, followed by co-incubation with CFSE-labeled OT-I CD8+ T cells for 64 hours. The number of CD8+ T cells that had undergone division as a measure of cross presentation was evaluated by flow cytometry. ( E ) DHFR activity of recombinant β-lactamase-DHFR fusion protein was measured after incubation with 500 nM MTX or equivalent volume PBS. ( F ) The ability of increasing amounts of recombinant β-lactamase-DHFR fusion protein to undergo ECT was evaluated in WT BMDCs following treatment with 500 nM MTX or equivalent volume PBS. *P

    Techniques Used: Incubation, Labeling, Flow Cytometry, Functional Assay, Activity Assay, Recombinant

    AQP3 is involved in cross presentation efficiency in vitro and in vivo . ( A ) ECT was evaluated in AQP3 -/- or WT BMDCs with 50 μg/ml β-lactamase. ( B-C ) Splenic CD11c+ DCs from AQP3 -/- or WT mice were isolated and ECT was evaluated in the CD11c+CD8+ population ( B ) or the CD11c+XCR1+ population ( C ) with 0.5 mg/ml β-lactamase. ( D ) BMDCs overexpressing AQP3 by viral transduction were incubated with increasing amounts of OVA or SIINFEKL peptide followed by co-incubation with CFSE-labeled OT-I CD8+ T cells. ( E ) BMDCs overexpressing AQP3 by viral transduction were incubated with 1 μg/ml αDEC205-OVA followed by co-incubation with CFSE-labeled OT-I CD8+ T cells. ( F ) BMDCs were incubated with 250 μg/ml OVA followed by co-incubation with CFSE-labeled OT-II CD4+ T cells. ( G ) ECT was evaluated in BMDCs expressing an AQP3 shRNA knockdown construct by viral transduction with 100 μg/ml β-lactamase. AQP3 shRNA knockdown was calculated to be 93% effective as measured by real-time PCR. ( H ) AQP3 KD cells were incubated with 1 μg/ml αDEC205-OVA followed by co-incubation with CFSE-labeled OT-I CD8+ T cells. ( I,J ) AQP3 -/- and WT control mice were infected with LCMV Armstrong. 8 days later, viral titer in kidney was assessed ( I ) and splenocytes were stimulated with LCMV NP peptide to determine the generation of LCMV-specific CD8+ T cell clones ( J ). Each dot represents an individual mouse. ( K ) AQP3 -/- and WT control mice were injected with rat HER-2/neu extended peptide and adjuvant. 1 week later, splenocytes were harvested and the generation of antigen-specific CD8+ T cell clones was evaluated using a HER-2/neu tetramer. Mean ± SEM is shown. *P
    Figure Legend Snippet: AQP3 is involved in cross presentation efficiency in vitro and in vivo . ( A ) ECT was evaluated in AQP3 -/- or WT BMDCs with 50 μg/ml β-lactamase. ( B-C ) Splenic CD11c+ DCs from AQP3 -/- or WT mice were isolated and ECT was evaluated in the CD11c+CD8+ population ( B ) or the CD11c+XCR1+ population ( C ) with 0.5 mg/ml β-lactamase. ( D ) BMDCs overexpressing AQP3 by viral transduction were incubated with increasing amounts of OVA or SIINFEKL peptide followed by co-incubation with CFSE-labeled OT-I CD8+ T cells. ( E ) BMDCs overexpressing AQP3 by viral transduction were incubated with 1 μg/ml αDEC205-OVA followed by co-incubation with CFSE-labeled OT-I CD8+ T cells. ( F ) BMDCs were incubated with 250 μg/ml OVA followed by co-incubation with CFSE-labeled OT-II CD4+ T cells. ( G ) ECT was evaluated in BMDCs expressing an AQP3 shRNA knockdown construct by viral transduction with 100 μg/ml β-lactamase. AQP3 shRNA knockdown was calculated to be 93% effective as measured by real-time PCR. ( H ) AQP3 KD cells were incubated with 1 μg/ml αDEC205-OVA followed by co-incubation with CFSE-labeled OT-I CD8+ T cells. ( I,J ) AQP3 -/- and WT control mice were infected with LCMV Armstrong. 8 days later, viral titer in kidney was assessed ( I ) and splenocytes were stimulated with LCMV NP peptide to determine the generation of LCMV-specific CD8+ T cell clones ( J ). Each dot represents an individual mouse. ( K ) AQP3 -/- and WT control mice were injected with rat HER-2/neu extended peptide and adjuvant. 1 week later, splenocytes were harvested and the generation of antigen-specific CD8+ T cell clones was evaluated using a HER-2/neu tetramer. Mean ± SEM is shown. *P

    Techniques Used: In Vitro, In Vivo, Mouse Assay, Isolation, Transduction, Incubation, Labeling, Expressing, shRNA, Construct, Real-time Polymerase Chain Reaction, Infection, Clone Assay, Injection

    AQP3 is uniquely positioned among aquaporin family members to regulate ECT. ( A ) HEK293 cells were transfected with various aquaporin overexpression constructs, flow cytometry-sorted for equivalent expression, and ECT efficiency (described in Methods) was evaluated with 100 μg/ml β-lactamase. Mean ± SEM is shown. ( B ) HEK293 cells were co-transfected with fluorescently-tagged AQP3 and AQP9 expression constructs and 3 days later, fixed and imaged by confocal microscopy. Scale bar: 2 μm. ( C ) HEK293 cells stably expressing fluorescently-tagged AQP3 were incubated with 1 μm latex beads for 2 hours prior to fixation and analysis by immuno-electron microscopy. 3 representative images are shown. Arrows point to phagosomal membrane. E: Endosome. P: Phagosome/latex bead. M: Mitochondria. Scale bar for top left image: 500 nm. Scale bar for top right and bottom left images: 200 nm. (D) On day 6 of culture, WT or AQP3 -/- BMDCS were fixed in 4% PFA and permeabilized with 0.05% saponin, followed by detection with anti-AQP3. AQP3 can be seen on or near the plasma membrane and dotting the interior of the cell. Scale bar: 2 μm. ( E ) HEK293 cells were transfected with fluorescently-tagged AQP3 or AQP3 2xLLmut. 3 days later, cells were incubated with fluorescently-labeled OVA-coated 1 μm beads for 1 hour prior to cell homogenization and analysis by flow cytometry. (F) AQP3 2xLLmut does not localize to phagosomes. Stably-transfected HEK293s were incubated with 100:1 (beads:cells) ratio of fluorescently-labeled OVA-coated 1 um tosylactivated Dynabeads and live imaging was initiated immediately. Scale bar: 1 μm. 2–4 independent experiments were performed.
    Figure Legend Snippet: AQP3 is uniquely positioned among aquaporin family members to regulate ECT. ( A ) HEK293 cells were transfected with various aquaporin overexpression constructs, flow cytometry-sorted for equivalent expression, and ECT efficiency (described in Methods) was evaluated with 100 μg/ml β-lactamase. Mean ± SEM is shown. ( B ) HEK293 cells were co-transfected with fluorescently-tagged AQP3 and AQP9 expression constructs and 3 days later, fixed and imaged by confocal microscopy. Scale bar: 2 μm. ( C ) HEK293 cells stably expressing fluorescently-tagged AQP3 were incubated with 1 μm latex beads for 2 hours prior to fixation and analysis by immuno-electron microscopy. 3 representative images are shown. Arrows point to phagosomal membrane. E: Endosome. P: Phagosome/latex bead. M: Mitochondria. Scale bar for top left image: 500 nm. Scale bar for top right and bottom left images: 200 nm. (D) On day 6 of culture, WT or AQP3 -/- BMDCS were fixed in 4% PFA and permeabilized with 0.05% saponin, followed by detection with anti-AQP3. AQP3 can be seen on or near the plasma membrane and dotting the interior of the cell. Scale bar: 2 μm. ( E ) HEK293 cells were transfected with fluorescently-tagged AQP3 or AQP3 2xLLmut. 3 days later, cells were incubated with fluorescently-labeled OVA-coated 1 μm beads for 1 hour prior to cell homogenization and analysis by flow cytometry. (F) AQP3 2xLLmut does not localize to phagosomes. Stably-transfected HEK293s were incubated with 100:1 (beads:cells) ratio of fluorescently-labeled OVA-coated 1 um tosylactivated Dynabeads and live imaging was initiated immediately. Scale bar: 1 μm. 2–4 independent experiments were performed.

    Techniques Used: Transfection, Over Expression, Construct, Flow Cytometry, Expressing, Confocal Microscopy, Stable Transfection, Incubation, Immuno-Electron Microscopy, Labeling, Homogenization, Imaging

    Two independent functional genomics screens for regulators of ECT uncover AQP3 . ( A ) CCF4 ECT assay schematic. ( B ) Representative flow cytometry plot of the sorting of WT BMDCs based on ECT+ or ECT- populations. ( C ) AQP3 was significantly enriched in the ECT+ population (false discovery rate adjusted p-value
    Figure Legend Snippet: Two independent functional genomics screens for regulators of ECT uncover AQP3 . ( A ) CCF4 ECT assay schematic. ( B ) Representative flow cytometry plot of the sorting of WT BMDCs based on ECT+ or ECT- populations. ( C ) AQP3 was significantly enriched in the ECT+ population (false discovery rate adjusted p-value

    Techniques Used: Functional Assay, Flow Cytometry

    5) Product Images from "Aquaporin-3 regulates endosome-to-cytosol transfer via lipid peroxidation for cross presentation"

    Article Title: Aquaporin-3 regulates endosome-to-cytosol transfer via lipid peroxidation for cross presentation

    Journal: bioRxiv

    doi: 10.1101/2020.08.19.256966

    Two independent functional genomics screens for regulators of ECT uncover AQP3 . ( A ) CCF4 ECT assay schematic. ( B ) Representative flow cytometry plot of the sorting of WT BMDCs based on ECT+ or ECTpopulations. ( C ) AQP3 was significantly enriched in the ECT+ population (false discovery rate adjusted p-value
    Figure Legend Snippet: Two independent functional genomics screens for regulators of ECT uncover AQP3 . ( A ) CCF4 ECT assay schematic. ( B ) Representative flow cytometry plot of the sorting of WT BMDCs based on ECT+ or ECTpopulations. ( C ) AQP3 was significantly enriched in the ECT+ population (false discovery rate adjusted p-value

    Techniques Used: Functional Assay, Flow Cytometry

    AQP3 is uniquely positioned among aquaporin family members to regulate ECT. ( A ) HEK293 cells were transfected with various aquaporin overexpression constructs, flow cytometry-sorted for equivalent expression, and ECT efficiency (described in Methods) was evaluated with 100 μg/ml β-lactamase. ( B ) HEK293 cells were co-transfected with fluorescently-tagged AQP3 and AQP9 expression constructs and 3 days later, fixed and imaged by confocal microscopy. Scale bar: 2 μm. ( C ) HEK293 cells stably expressing fluorescently-tagged AQP3 were incubated with 1 μm latex beads for 2 hours prior to fixation and analysis by immuno-electron microscopy. 3 representative images are shown. Arrows point to phagosomal membrane. E: endosome. P: phagosome/latex bead. M: mitochondria. Scale bar for top left image: 500 nm. Scale bar for top right and bottom left images: 200 nm. ( D ) On day 6 of culture, WT or AQP3 -/- BMDCS were fixed in 4% PFA and permeabilized with 0.05% saponin, followed by detection with anti-AQP3. AQP3 can be seen on or near the plasma membrane and dotting the interior of the cell. Scale bar: 2 μm. ( E ) HEK293 cells were transfected with fluorescently-tagged AQP3 or AQP3 2xLLmut. 3 days later, cells were incubated with fluorescently-labeled OVA-coated 1 μm beads for 1 hour prior to cell homogenization and analysis by flow cytometry. ( F ) AQP3 2xLLmut does not localize to phagosomes. Stably-transfected HEK293s were incubated with 100:1 (beads:cells) ratio of fluorescently-labeled OVA-coated 1 um tosylactivated Dynabeads and live imaging was initiated immediately. Scale bar: 1 μm.
    Figure Legend Snippet: AQP3 is uniquely positioned among aquaporin family members to regulate ECT. ( A ) HEK293 cells were transfected with various aquaporin overexpression constructs, flow cytometry-sorted for equivalent expression, and ECT efficiency (described in Methods) was evaluated with 100 μg/ml β-lactamase. ( B ) HEK293 cells were co-transfected with fluorescently-tagged AQP3 and AQP9 expression constructs and 3 days later, fixed and imaged by confocal microscopy. Scale bar: 2 μm. ( C ) HEK293 cells stably expressing fluorescently-tagged AQP3 were incubated with 1 μm latex beads for 2 hours prior to fixation and analysis by immuno-electron microscopy. 3 representative images are shown. Arrows point to phagosomal membrane. E: endosome. P: phagosome/latex bead. M: mitochondria. Scale bar for top left image: 500 nm. Scale bar for top right and bottom left images: 200 nm. ( D ) On day 6 of culture, WT or AQP3 -/- BMDCS were fixed in 4% PFA and permeabilized with 0.05% saponin, followed by detection with anti-AQP3. AQP3 can be seen on or near the plasma membrane and dotting the interior of the cell. Scale bar: 2 μm. ( E ) HEK293 cells were transfected with fluorescently-tagged AQP3 or AQP3 2xLLmut. 3 days later, cells were incubated with fluorescently-labeled OVA-coated 1 μm beads for 1 hour prior to cell homogenization and analysis by flow cytometry. ( F ) AQP3 2xLLmut does not localize to phagosomes. Stably-transfected HEK293s were incubated with 100:1 (beads:cells) ratio of fluorescently-labeled OVA-coated 1 um tosylactivated Dynabeads and live imaging was initiated immediately. Scale bar: 1 μm.

    Techniques Used: Transfection, Over Expression, Construct, Flow Cytometry, Expressing, Confocal Microscopy, Stable Transfection, Incubation, Immuno-Electron Microscopy, Labeling, Homogenization, Imaging

    AQP3 is involved in cross presentation efficiency in vitro and in vivo . ( A ) ECT was evaluated in AQP3 -/- or WT BMDCs with 50 μg/ml β-lactamase. ( B-C ) Splenic CD11c+ DCs from AQP3 -/- or WT mice were isolated and ECT was evaluated in the CD11c+CD8+ population ( B ) or the CD11c+XCR1+ population ( C ) with 0.5 mg/ml β-lactamase. ( D ) BMDCs overexpressing AQP3 by viral transduction were incubated with increasing amounts of OVA or SIINFEKL peptide followed by co-incubation with CFSE-labeled OT-I CD8+ T cells. ( E ) BMDCs overexpressing AQP3 by viral transduction were incubated with 1 μg/ml αDEC205-OVA followed by co-incubation with CFSE-labeled OT-I CD8+ T cells. ( F ) BMDCs were incubated with 250 μg/ml OVA followed by coincubation with CFSE-labeled OT-II CD4+ T cells. ( G ) ECT was evaluated in BMDCs expressing an AQP3 shRNA knockdown construct by viral transduction with 100 μg/ml β-lactamase. AQP3 shRNA knockdown was calculated to be 93% effective as measured by real-time PCR. ( H ) AQP3 KD cells were incubated with 1 μg/ml αDEC205-OVA followed by co-incubation with CFSE-labeled OT-I CD8+ T cells. ( I,J ) AQP3 -/- and WT control mice were infected with LCMV Armstrong. 8 days later, viral titer in kidney was assessed ( I ) and splenocytes were stimulated with LCMV NP peptide to determine the generation of LCMV-specific CD8+ T cell clones (J). Each dot represents an individual mouse. ( K ) AQP3 -/- and WT control mice were injected with rat HER-2/neu extended peptide and adjuvant. 1 week later, splenocytes were harvested and the generation of antigen-specific CD8+ T cell clones was evaluated using a HER-2/neu tetramer. *P
    Figure Legend Snippet: AQP3 is involved in cross presentation efficiency in vitro and in vivo . ( A ) ECT was evaluated in AQP3 -/- or WT BMDCs with 50 μg/ml β-lactamase. ( B-C ) Splenic CD11c+ DCs from AQP3 -/- or WT mice were isolated and ECT was evaluated in the CD11c+CD8+ population ( B ) or the CD11c+XCR1+ population ( C ) with 0.5 mg/ml β-lactamase. ( D ) BMDCs overexpressing AQP3 by viral transduction were incubated with increasing amounts of OVA or SIINFEKL peptide followed by co-incubation with CFSE-labeled OT-I CD8+ T cells. ( E ) BMDCs overexpressing AQP3 by viral transduction were incubated with 1 μg/ml αDEC205-OVA followed by co-incubation with CFSE-labeled OT-I CD8+ T cells. ( F ) BMDCs were incubated with 250 μg/ml OVA followed by coincubation with CFSE-labeled OT-II CD4+ T cells. ( G ) ECT was evaluated in BMDCs expressing an AQP3 shRNA knockdown construct by viral transduction with 100 μg/ml β-lactamase. AQP3 shRNA knockdown was calculated to be 93% effective as measured by real-time PCR. ( H ) AQP3 KD cells were incubated with 1 μg/ml αDEC205-OVA followed by co-incubation with CFSE-labeled OT-I CD8+ T cells. ( I,J ) AQP3 -/- and WT control mice were infected with LCMV Armstrong. 8 days later, viral titer in kidney was assessed ( I ) and splenocytes were stimulated with LCMV NP peptide to determine the generation of LCMV-specific CD8+ T cell clones (J). Each dot represents an individual mouse. ( K ) AQP3 -/- and WT control mice were injected with rat HER-2/neu extended peptide and adjuvant. 1 week later, splenocytes were harvested and the generation of antigen-specific CD8+ T cell clones was evaluated using a HER-2/neu tetramer. *P

    Techniques Used: In Vitro, In Vivo, Mouse Assay, Isolation, Transduction, Incubation, Labeling, Expressing, shRNA, Construct, Real-time Polymerase Chain Reaction, Infection, Injection, Clone Assay

    AQP3 regulates phagosomal lipid peroxidation. ( A ) AQP3 -/- or WT BMDCs were co-incubated with fluorescently-labeled OVA-coated 1 μm beads and a C11-bodipy lipid peroxidation indicator for 1.5 hours, homogenized, and phagosomes were analyzed by flow cytometry. ( B ) Annotated volcano plot of ECT functional genomics screen highlights the connection between ECT and ROS generation and transport, which could potentiate endosomal lipid peroxidation and antigen release. ( C ) ECT was evaluated in WT BMDCs with 0.5 mg/ml β-lactamase in media that did or did not contain ISCOMATRIX™ adjuvant. ( D ) WT BMDCs were incubated with 100 mg/ml OVA in media that did or did not contain ISCOMATRIX™ adjuvant, in the presence or absence of 1 μ M epoxomicin, followed by co-incubation with CFSE-labeled OT-I CD8+ T cells. The number of CD8+ T cells that had undergone division as a measure of cross presentation was evaluated by flow cytometry. ( E ) DHFR activity of recombinant β-lactamase-DHFR fusion protein was measured after incubation with 500 nM MTX or equivalent volume PBS. ( F ) The ability of increasing amounts of recombinant β-lactamase-DHFR fusion protein to undergo ECT was evaluated in WT BMDCs following treatment with 500 nM MTX or equivalent volume PBS. *P
    Figure Legend Snippet: AQP3 regulates phagosomal lipid peroxidation. ( A ) AQP3 -/- or WT BMDCs were co-incubated with fluorescently-labeled OVA-coated 1 μm beads and a C11-bodipy lipid peroxidation indicator for 1.5 hours, homogenized, and phagosomes were analyzed by flow cytometry. ( B ) Annotated volcano plot of ECT functional genomics screen highlights the connection between ECT and ROS generation and transport, which could potentiate endosomal lipid peroxidation and antigen release. ( C ) ECT was evaluated in WT BMDCs with 0.5 mg/ml β-lactamase in media that did or did not contain ISCOMATRIX™ adjuvant. ( D ) WT BMDCs were incubated with 100 mg/ml OVA in media that did or did not contain ISCOMATRIX™ adjuvant, in the presence or absence of 1 μ M epoxomicin, followed by co-incubation with CFSE-labeled OT-I CD8+ T cells. The number of CD8+ T cells that had undergone division as a measure of cross presentation was evaluated by flow cytometry. ( E ) DHFR activity of recombinant β-lactamase-DHFR fusion protein was measured after incubation with 500 nM MTX or equivalent volume PBS. ( F ) The ability of increasing amounts of recombinant β-lactamase-DHFR fusion protein to undergo ECT was evaluated in WT BMDCs following treatment with 500 nM MTX or equivalent volume PBS. *P

    Techniques Used: Incubation, Labeling, Flow Cytometry, Functional Assay, Activity Assay, Recombinant

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    Alomone Labs antibody against aqp3
    A549 cell aggregation is an active process. ( A ) The effect of ROCK on A549 cell aggregation. The phase-contrast micrographs show the morphologies of the A549 cells following treatment with a ROCK inhibitor, Y-27632, and the Rho activator Ⅱ. ( B ) The effects of <t>AQP3</t> knockdown with siRNA on actomyosin cytoskeleton remodeling. A549 cells were stained with anti-AQP3 antibody, followed by Fluorescein-conjugated antibody (green). The actin microfilaments were stained with rhodamine-conjugated phalloidin (red), and the nuclei were stained with DAPI (blue). ( C ) The effects of AQP3 knockdown with siRNA on the apoptosis signaling pathway. Twenty-four h following siRNA transfection in 2D culture condition, the cells were further incubated in the 2D or 3D culture condition. Then, cells were harvested to confirm the effect of AQP3 knockdown on the apoptotic molecular signatures using Western blotting. α-tubulin was used as an internal control.
    Antibody Against Aqp3, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Alomone Labs anti aqp3
    <t>AQP3</t> Immunoreactivity was Mislocalized in a Majority of Psoriatic Lesions (A through C) Archived paraffin-embedded psoriasis samples were sectioned (4 μm) and deparaffinized. Sections were then stained with an antibody recognizing AQP3 and visualized with an ABC kit and DAB. Results from three patients are shown and the staining pattern is representative of 8 of 10 psoriatic samples examined. (D) A negative control was performed by omitting primary antibody. Note that AQP3 is localized to the cytoplasm rather than to the plasma membrane in a majority of the psoriatic epidermis. All sections were counterstained with hematoxylin.
    Anti Aqp3, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Alomone Labs antibody recognizing aqp3
    <t>AQP3</t> and PLD2 co-localize in the cornea in situ and in immortalized human corneal limbal epithelial cells in vitro. Formalin-fixed human corneas obtained from the Georgia Eye Bank were paraffin-embedded and sectioned onto microscope slides. Sections were stained using an Opal immunofluorescence kit and incubated with a rabbit polyclonal antibody recognizing PLD2 and a mouse monoclonal antibody against AQP3. Sections were then processed according to the supplier's instructions with red staining (Opal 570) representing PLD2 and green (Opal 520) representing AQP3. Staining was visualized using confocal microscopy on a Zeiss laser-scanning microscope. ( A ) A more central region of the corneal epithelium, ( B ) the limbal region of the cornea, and ( C ) immortalized human corneal limbal epithelial cells are shown. Results are representative of at least three corneas or cell passages; negative controls in which primary antibodies were omitted demonstrated essentially no staining.
    Antibody Recognizing Aqp3, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 92/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Alomone Labs rabbit anti rat aquaporin 3 aqp3 antibody
    Expression levels and distribution of aquaporin (AQP) in mouse skin. Erlotinib was administered orally to the mice for 14 days, and the AQP (AQP0-9) mRNA expression level in the skin was measured by real-time RT-PCR. After normalization to 18S rRNA, the data are presented with the mean value of the control group set at 100% ( A ). The protein expression of <t>AQP3</t> in the skin was analyzed by Western blotting. After normalization to glyceraldehyde-3-phosphate dehydrogenase (GAPDH), the data are shown with the mean value of the control group set at 100% (mean ± SD, n = 5; * p
    Rabbit Anti Rat Aquaporin 3 Aqp3 Antibody, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 85/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    A549 cell aggregation is an active process. ( A ) The effect of ROCK on A549 cell aggregation. The phase-contrast micrographs show the morphologies of the A549 cells following treatment with a ROCK inhibitor, Y-27632, and the Rho activator Ⅱ. ( B ) The effects of AQP3 knockdown with siRNA on actomyosin cytoskeleton remodeling. A549 cells were stained with anti-AQP3 antibody, followed by Fluorescein-conjugated antibody (green). The actin microfilaments were stained with rhodamine-conjugated phalloidin (red), and the nuclei were stained with DAPI (blue). ( C ) The effects of AQP3 knockdown with siRNA on the apoptosis signaling pathway. Twenty-four h following siRNA transfection in 2D culture condition, the cells were further incubated in the 2D or 3D culture condition. Then, cells were harvested to confirm the effect of AQP3 knockdown on the apoptotic molecular signatures using Western blotting. α-tubulin was used as an internal control.

    Journal: International Journal of Molecular Sciences

    Article Title: AQP3 Increases Intercellular Cohesion in NSCLC A549 Cell Spheroids through Exploratory Cell Protrusions

    doi: 10.3390/ijms22084287

    Figure Lengend Snippet: A549 cell aggregation is an active process. ( A ) The effect of ROCK on A549 cell aggregation. The phase-contrast micrographs show the morphologies of the A549 cells following treatment with a ROCK inhibitor, Y-27632, and the Rho activator Ⅱ. ( B ) The effects of AQP3 knockdown with siRNA on actomyosin cytoskeleton remodeling. A549 cells were stained with anti-AQP3 antibody, followed by Fluorescein-conjugated antibody (green). The actin microfilaments were stained with rhodamine-conjugated phalloidin (red), and the nuclei were stained with DAPI (blue). ( C ) The effects of AQP3 knockdown with siRNA on the apoptosis signaling pathway. Twenty-four h following siRNA transfection in 2D culture condition, the cells were further incubated in the 2D or 3D culture condition. Then, cells were harvested to confirm the effect of AQP3 knockdown on the apoptotic molecular signatures using Western blotting. α-tubulin was used as an internal control.

    Article Snippet: The samples were then separated with SDS-PAGE gel and immunoblotted with the antibody against AQP3 (Alomone Labs Ltd., Jerusalem, Israel, 1/200), GAPDH (BioLegend, San Diego, CA, USA), or β-actin (Santa Cruz Biotechnology) or α-tubulin (Santa Cruz Biotechnology). β-actin, GAPDH, and α-tubulin were used as loading controls.

    Techniques: Staining, Transfection, Incubation, Western Blot

    The effect of AQP3 on spatiotemporal dynamics of protrusions. ( A ) Polymerase chain reaction and ( B ) Quantitative real-time reverse transcription-polymerase chain reaction of the transcript levels of the organic hydroxyl transport genes. The data shown here represent three independent experiments, and the values represent the mean ± SEM of triplicate samples. The level of each mRNA was normalized to that of the GAPDH mRNA in the same sample and presented as the fold-change over that of the 2D culture control cells. The differences in expression levels were evaluated for significance using two-tailed t-tests with unequal variance. * p

    Journal: International Journal of Molecular Sciences

    Article Title: AQP3 Increases Intercellular Cohesion in NSCLC A549 Cell Spheroids through Exploratory Cell Protrusions

    doi: 10.3390/ijms22084287

    Figure Lengend Snippet: The effect of AQP3 on spatiotemporal dynamics of protrusions. ( A ) Polymerase chain reaction and ( B ) Quantitative real-time reverse transcription-polymerase chain reaction of the transcript levels of the organic hydroxyl transport genes. The data shown here represent three independent experiments, and the values represent the mean ± SEM of triplicate samples. The level of each mRNA was normalized to that of the GAPDH mRNA in the same sample and presented as the fold-change over that of the 2D culture control cells. The differences in expression levels were evaluated for significance using two-tailed t-tests with unequal variance. * p

    Article Snippet: The samples were then separated with SDS-PAGE gel and immunoblotted with the antibody against AQP3 (Alomone Labs Ltd., Jerusalem, Israel, 1/200), GAPDH (BioLegend, San Diego, CA, USA), or β-actin (Santa Cruz Biotechnology) or α-tubulin (Santa Cruz Biotechnology). β-actin, GAPDH, and α-tubulin were used as loading controls.

    Techniques: Polymerase Chain Reaction, Reverse Transcription Polymerase Chain Reaction, Expressing, Two Tailed Test

    AQP3 Immunoreactivity was Mislocalized in a Majority of Psoriatic Lesions (A through C) Archived paraffin-embedded psoriasis samples were sectioned (4 μm) and deparaffinized. Sections were then stained with an antibody recognizing AQP3 and visualized with an ABC kit and DAB. Results from three patients are shown and the staining pattern is representative of 8 of 10 psoriatic samples examined. (D) A negative control was performed by omitting primary antibody. Note that AQP3 is localized to the cytoplasm rather than to the plasma membrane in a majority of the psoriatic epidermis. All sections were counterstained with hematoxylin.

    Journal: Archives of dermatological research

    Article Title: ABNORMAL AQUAPORIN-3 PROTEIN EXPRESSION IN HYPERPROLIFERATIVE SKIN DISORDERS

    doi: 10.1007/s00403-011-1136-x

    Figure Lengend Snippet: AQP3 Immunoreactivity was Mislocalized in a Majority of Psoriatic Lesions (A through C) Archived paraffin-embedded psoriasis samples were sectioned (4 μm) and deparaffinized. Sections were then stained with an antibody recognizing AQP3 and visualized with an ABC kit and DAB. Results from three patients are shown and the staining pattern is representative of 8 of 10 psoriatic samples examined. (D) A negative control was performed by omitting primary antibody. Note that AQP3 is localized to the cytoplasm rather than to the plasma membrane in a majority of the psoriatic epidermis. All sections were counterstained with hematoxylin.

    Article Snippet: Slides for AQP3 staining were deparaffinized, washed twice for 5 minutes in phosphate-buffered saline (PBS), incubated 30 minutes in 3% hydrogen peroxide, washed twice for 5 minutes in PBS, incubated 1 hour in 0.3% goat serum, and then incubated overnight with anti-AQP3 (Alomone Labs, Jerusalem, Israel; 1:1000 dilution) in a humidified chamber at 4°C.

    Techniques: Staining, Negative Control

    AQP3 Immunoreactivity was “Patchy” in Squamous Cell Carcinoma (SCC) Archived paraffin-embedded squamous cell carcinoma (SCC) samples were sectioned (4 μm) and deparaffinized. Sections were then stained with antibodies recognizing AQP3 and visualized with an ABC kit and DAB. Sections were counterstained with hematoxylin. Illustrated in panels A through C is AQP3 staining in 3 patients with results representative of 5 of 5 SCCs. A negative control in which the primary antibody was omitted showed no AQP3 staining (panel D).

    Journal: Archives of dermatological research

    Article Title: ABNORMAL AQUAPORIN-3 PROTEIN EXPRESSION IN HYPERPROLIFERATIVE SKIN DISORDERS

    doi: 10.1007/s00403-011-1136-x

    Figure Lengend Snippet: AQP3 Immunoreactivity was “Patchy” in Squamous Cell Carcinoma (SCC) Archived paraffin-embedded squamous cell carcinoma (SCC) samples were sectioned (4 μm) and deparaffinized. Sections were then stained with antibodies recognizing AQP3 and visualized with an ABC kit and DAB. Sections were counterstained with hematoxylin. Illustrated in panels A through C is AQP3 staining in 3 patients with results representative of 5 of 5 SCCs. A negative control in which the primary antibody was omitted showed no AQP3 staining (panel D).

    Article Snippet: Slides for AQP3 staining were deparaffinized, washed twice for 5 minutes in phosphate-buffered saline (PBS), incubated 30 minutes in 3% hydrogen peroxide, washed twice for 5 minutes in PBS, incubated 1 hour in 0.3% goat serum, and then incubated overnight with anti-AQP3 (Alomone Labs, Jerusalem, Israel; 1:1000 dilution) in a humidified chamber at 4°C.

    Techniques: Staining, Negative Control

    AQP3 Staining was Reduced in Ki67-Positive Cells in Squamous Cell Carcinoma Archived paraffin-embedded squamous cell carcinoma (SCC) samples were sectioned (4 μm) and deparaffinized. Sequential serial sections were then stained with antibodies recognizing (A, C) AQP3 or (B, D) Ki67 [an antigen expressed in proliferating cells (Dako, Carpinteria, CA)] and visualized with an ABC kit and DAB. Sections were counterstained with hematoxylin. Note that cells that are Ki67 positive exhibit reduced or absent AQP3 staining (arrows in panels C and D). The inverse correlation of Ki67 positivity with AQP3 immunoreactivity is representative of 4 of 4 SCCs. (E) Human tonsil stained with Ki67 as a positive control. (F) Primary antibody was omitted as a negative control.

    Journal: Archives of dermatological research

    Article Title: ABNORMAL AQUAPORIN-3 PROTEIN EXPRESSION IN HYPERPROLIFERATIVE SKIN DISORDERS

    doi: 10.1007/s00403-011-1136-x

    Figure Lengend Snippet: AQP3 Staining was Reduced in Ki67-Positive Cells in Squamous Cell Carcinoma Archived paraffin-embedded squamous cell carcinoma (SCC) samples were sectioned (4 μm) and deparaffinized. Sequential serial sections were then stained with antibodies recognizing (A, C) AQP3 or (B, D) Ki67 [an antigen expressed in proliferating cells (Dako, Carpinteria, CA)] and visualized with an ABC kit and DAB. Sections were counterstained with hematoxylin. Note that cells that are Ki67 positive exhibit reduced or absent AQP3 staining (arrows in panels C and D). The inverse correlation of Ki67 positivity with AQP3 immunoreactivity is representative of 4 of 4 SCCs. (E) Human tonsil stained with Ki67 as a positive control. (F) Primary antibody was omitted as a negative control.

    Article Snippet: Slides for AQP3 staining were deparaffinized, washed twice for 5 minutes in phosphate-buffered saline (PBS), incubated 30 minutes in 3% hydrogen peroxide, washed twice for 5 minutes in PBS, incubated 1 hour in 0.3% goat serum, and then incubated overnight with anti-AQP3 (Alomone Labs, Jerusalem, Israel; 1:1000 dilution) in a humidified chamber at 4°C.

    Techniques: Staining, Positive Control, Negative Control

    AQP3 Immunoreactivity was Reduced or Absent in Basal Cell Carcinoma (BCC) Archived paraffin-embedded basal cell carcinoma samples were sectioned (4 μm), deparaffinized, stained with antibodies recognizing AQP3 and visualized with an ABC kit and 3,3’-diaminobenzidine (DAB). Sections were counterstained with hematoxylin. Results illustrate three BCCs from different patients and are representative of 13 of 13 BCCs. The results were quantified as 3+ (intense staining), 2+ (moderate staining), + (weak staining) or 0 (no staining) by three independent observers and the values averaged. Shown in panel D is the quantitation (means ± SEM) of the 13 BCCs relative to normal-appearing overlying epidermis with the data analyzed using a student's t-test; *p

    Journal: Archives of dermatological research

    Article Title: ABNORMAL AQUAPORIN-3 PROTEIN EXPRESSION IN HYPERPROLIFERATIVE SKIN DISORDERS

    doi: 10.1007/s00403-011-1136-x

    Figure Lengend Snippet: AQP3 Immunoreactivity was Reduced or Absent in Basal Cell Carcinoma (BCC) Archived paraffin-embedded basal cell carcinoma samples were sectioned (4 μm), deparaffinized, stained with antibodies recognizing AQP3 and visualized with an ABC kit and 3,3’-diaminobenzidine (DAB). Sections were counterstained with hematoxylin. Results illustrate three BCCs from different patients and are representative of 13 of 13 BCCs. The results were quantified as 3+ (intense staining), 2+ (moderate staining), + (weak staining) or 0 (no staining) by three independent observers and the values averaged. Shown in panel D is the quantitation (means ± SEM) of the 13 BCCs relative to normal-appearing overlying epidermis with the data analyzed using a student's t-test; *p

    Article Snippet: Slides for AQP3 staining were deparaffinized, washed twice for 5 minutes in phosphate-buffered saline (PBS), incubated 30 minutes in 3% hydrogen peroxide, washed twice for 5 minutes in PBS, incubated 1 hour in 0.3% goat serum, and then incubated overnight with anti-AQP3 (Alomone Labs, Jerusalem, Israel; 1:1000 dilution) in a humidified chamber at 4°C.

    Techniques: Staining, Quantitation Assay

    AQP3 and PLD2 co-localize in the cornea in situ and in immortalized human corneal limbal epithelial cells in vitro. Formalin-fixed human corneas obtained from the Georgia Eye Bank were paraffin-embedded and sectioned onto microscope slides. Sections were stained using an Opal immunofluorescence kit and incubated with a rabbit polyclonal antibody recognizing PLD2 and a mouse monoclonal antibody against AQP3. Sections were then processed according to the supplier's instructions with red staining (Opal 570) representing PLD2 and green (Opal 520) representing AQP3. Staining was visualized using confocal microscopy on a Zeiss laser-scanning microscope. ( A ) A more central region of the corneal epithelium, ( B ) the limbal region of the cornea, and ( C ) immortalized human corneal limbal epithelial cells are shown. Results are representative of at least three corneas or cell passages; negative controls in which primary antibodies were omitted demonstrated essentially no staining.

    Journal: Investigative Ophthalmology & Visual Science

    Article Title: Dioleoylphosphatidylglycerol Accelerates Corneal Epithelial Wound Healing

    doi: 10.1167/iovs.61.3.29

    Figure Lengend Snippet: AQP3 and PLD2 co-localize in the cornea in situ and in immortalized human corneal limbal epithelial cells in vitro. Formalin-fixed human corneas obtained from the Georgia Eye Bank were paraffin-embedded and sectioned onto microscope slides. Sections were stained using an Opal immunofluorescence kit and incubated with a rabbit polyclonal antibody recognizing PLD2 and a mouse monoclonal antibody against AQP3. Sections were then processed according to the supplier's instructions with red staining (Opal 570) representing PLD2 and green (Opal 520) representing AQP3. Staining was visualized using confocal microscopy on a Zeiss laser-scanning microscope. ( A ) A more central region of the corneal epithelium, ( B ) the limbal region of the cornea, and ( C ) immortalized human corneal limbal epithelial cells are shown. Results are representative of at least three corneas or cell passages; negative controls in which primary antibodies were omitted demonstrated essentially no staining.

    Article Snippet: The antibody recognizing AQP3 (Alomone Labs, Jerusalem, Israel) was previously validated in our laboratory using western analysis and by demonstrating increased AQP3 immunoreactivity in the skin of an epidermal-targeted AQP3-overexpressing transgenic mouse model. , .

    Techniques: In Situ, In Vitro, Microscopy, Staining, Immunofluorescence, Incubation, Confocal Microscopy, Laser-Scanning Microscopy

    The ability of DOPG to enhance corneal epithelial wound healing is not affected by the sex of the animal. Male and female AQP3 knockout mice treated as in Figure 6 were analyzed separately (n = 4–6). Regression slopes (the healing rate) were compared by an unpaired Student's t -test and illustrated as values for individual animals by sex as indicated ( * P

    Journal: Investigative Ophthalmology & Visual Science

    Article Title: Dioleoylphosphatidylglycerol Accelerates Corneal Epithelial Wound Healing

    doi: 10.1167/iovs.61.3.29

    Figure Lengend Snippet: The ability of DOPG to enhance corneal epithelial wound healing is not affected by the sex of the animal. Male and female AQP3 knockout mice treated as in Figure 6 were analyzed separately (n = 4–6). Regression slopes (the healing rate) were compared by an unpaired Student's t -test and illustrated as values for individual animals by sex as indicated ( * P

    Article Snippet: The antibody recognizing AQP3 (Alomone Labs, Jerusalem, Israel) was previously validated in our laboratory using western analysis and by demonstrating increased AQP3 immunoreactivity in the skin of an epidermal-targeted AQP3-overexpressing transgenic mouse model. , .

    Techniques: Knock-Out, Mouse Assay

    Immortalized human corneal epithelial cells express both AQP3 and PLD2 proteins, which can be co-immunoprecipitated from these cells. SV40-immortalized human corneal epithelial cells were cultured in a 1:1 mixture of defined keratinocyte serum-free (DKSF) medium and Minimum Essential Medium until approximately 70% confluent. The cells were harvested using RIPA buffer and immunoprecipitated using antibody recognizing AQP3 as described in the Materials and Methods section. Immunoprecipitates were collected and analyzed by western blotting with antibodies recognizing AQP3 or PLD2 as indicated. Molecular weight standards were separated in the far right lane and appear as light bands. Results are representative of at least three separate experiments.

    Journal: Investigative Ophthalmology & Visual Science

    Article Title: Dioleoylphosphatidylglycerol Accelerates Corneal Epithelial Wound Healing

    doi: 10.1167/iovs.61.3.29

    Figure Lengend Snippet: Immortalized human corneal epithelial cells express both AQP3 and PLD2 proteins, which can be co-immunoprecipitated from these cells. SV40-immortalized human corneal epithelial cells were cultured in a 1:1 mixture of defined keratinocyte serum-free (DKSF) medium and Minimum Essential Medium until approximately 70% confluent. The cells were harvested using RIPA buffer and immunoprecipitated using antibody recognizing AQP3 as described in the Materials and Methods section. Immunoprecipitates were collected and analyzed by western blotting with antibodies recognizing AQP3 or PLD2 as indicated. Molecular weight standards were separated in the far right lane and appear as light bands. Results are representative of at least three separate experiments.

    Article Snippet: The antibody recognizing AQP3 (Alomone Labs, Jerusalem, Israel) was previously validated in our laboratory using western analysis and by demonstrating increased AQP3 immunoreactivity in the skin of an epidermal-targeted AQP3-overexpressing transgenic mouse model. , .

    Techniques: Immunoprecipitation, Cell Culture, Western Blot, Molecular Weight

    The interaction between PLD2 and AQP3 is likely direct. Sf9 insect cells were infected with baculovirus expressing either His-tagged PLD2 (PLD2-His; Fig. 4 A, left panel) or GST-tagged AQP3 (AQP3-GST; Fig. 4 B) alone or with both AQP3-GST and PLD2-His ( Fig. 4 A, right panel, Fig. 4 B). Equal volumes of pre-cleared Sf9 lysates were then immunoprecipitated (IP) with anti-GST antibody (to pull down AQP3-GST) or anti-PLD2 antibody, and the immunoprecipitates were analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblotting (IB) using antibodies recognizing GST (for AQP3-GST) or His (for PLD2-His) as indicated. A 1/10 volume of lysate was similarly analyzed (Input). Results are representative of three experiments.

    Journal: Investigative Ophthalmology & Visual Science

    Article Title: Dioleoylphosphatidylglycerol Accelerates Corneal Epithelial Wound Healing

    doi: 10.1167/iovs.61.3.29

    Figure Lengend Snippet: The interaction between PLD2 and AQP3 is likely direct. Sf9 insect cells were infected with baculovirus expressing either His-tagged PLD2 (PLD2-His; Fig. 4 A, left panel) or GST-tagged AQP3 (AQP3-GST; Fig. 4 B) alone or with both AQP3-GST and PLD2-His ( Fig. 4 A, right panel, Fig. 4 B). Equal volumes of pre-cleared Sf9 lysates were then immunoprecipitated (IP) with anti-GST antibody (to pull down AQP3-GST) or anti-PLD2 antibody, and the immunoprecipitates were analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblotting (IB) using antibodies recognizing GST (for AQP3-GST) or His (for PLD2-His) as indicated. A 1/10 volume of lysate was similarly analyzed (Input). Results are representative of three experiments.

    Article Snippet: The antibody recognizing AQP3 (Alomone Labs, Jerusalem, Israel) was previously validated in our laboratory using western analysis and by demonstrating increased AQP3 immunoreactivity in the skin of an epidermal-targeted AQP3-overexpressing transgenic mouse model. , .

    Techniques: Infection, Expressing, Immunoprecipitation, Polyacrylamide Gel Electrophoresis, SDS Page

    Human corneal epithelial cells express AQP3 in situ. Formalin-fixed human corneas obtained from the Georgia Eye Bank were paraffin-embedded and sectioned. After antigen retrieval, sections were incubated with an antibody recognizing AQP3 and visualized with an ABC staining kit using 3,3′-diaminobenzidine as chromogen (brown staining), as described in the Materials and Methods section. Sections were counterstained with hematoxylin (blue staining) and photographed. Results are representative of the staining of random sections of right and left corneas from at least three individuals. ( A , B ) AQP3 staining observed in two different corneas. ( C ) A negative control performed by omission of the primary antibody.

    Journal: Investigative Ophthalmology & Visual Science

    Article Title: Dioleoylphosphatidylglycerol Accelerates Corneal Epithelial Wound Healing

    doi: 10.1167/iovs.61.3.29

    Figure Lengend Snippet: Human corneal epithelial cells express AQP3 in situ. Formalin-fixed human corneas obtained from the Georgia Eye Bank were paraffin-embedded and sectioned. After antigen retrieval, sections were incubated with an antibody recognizing AQP3 and visualized with an ABC staining kit using 3,3′-diaminobenzidine as chromogen (brown staining), as described in the Materials and Methods section. Sections were counterstained with hematoxylin (blue staining) and photographed. Results are representative of the staining of random sections of right and left corneas from at least three individuals. ( A , B ) AQP3 staining observed in two different corneas. ( C ) A negative control performed by omission of the primary antibody.

    Article Snippet: The antibody recognizing AQP3 (Alomone Labs, Jerusalem, Israel) was previously validated in our laboratory using western analysis and by demonstrating increased AQP3 immunoreactivity in the skin of an epidermal-targeted AQP3-overexpressing transgenic mouse model. , .

    Techniques: In Situ, Incubation, Staining, Negative Control

    DOPG accelerates corneal epithelial wound healing in AQP3 knockout mice. Anesthetized mice (n = 10–12) received corneal wounds produced by scraping the epithelium with an Algerbrush. Fluorescein was added to the wounded eye, and the wound was photographed. Saline control vehicle (Con) or DOPG in saline (100 µg/mL) was then applied to the eye. This process was repeated every 4 hours for 28 hours. The area of the photographed wounds was digitized and expressed as a percentage of the initial wound area. ( A ) Mean ± SEM of the wound area at each Con versus DOPG time point; these values were compared using an unpaired Student's t-test, with ** P

    Journal: Investigative Ophthalmology & Visual Science

    Article Title: Dioleoylphosphatidylglycerol Accelerates Corneal Epithelial Wound Healing

    doi: 10.1167/iovs.61.3.29

    Figure Lengend Snippet: DOPG accelerates corneal epithelial wound healing in AQP3 knockout mice. Anesthetized mice (n = 10–12) received corneal wounds produced by scraping the epithelium with an Algerbrush. Fluorescein was added to the wounded eye, and the wound was photographed. Saline control vehicle (Con) or DOPG in saline (100 µg/mL) was then applied to the eye. This process was repeated every 4 hours for 28 hours. The area of the photographed wounds was digitized and expressed as a percentage of the initial wound area. ( A ) Mean ± SEM of the wound area at each Con versus DOPG time point; these values were compared using an unpaired Student's t-test, with ** P

    Article Snippet: The antibody recognizing AQP3 (Alomone Labs, Jerusalem, Israel) was previously validated in our laboratory using western analysis and by demonstrating increased AQP3 immunoreactivity in the skin of an epidermal-targeted AQP3-overexpressing transgenic mouse model. , .

    Techniques: Knock-Out, Mouse Assay, Produced

    Expression levels and distribution of aquaporin (AQP) in mouse skin. Erlotinib was administered orally to the mice for 14 days, and the AQP (AQP0-9) mRNA expression level in the skin was measured by real-time RT-PCR. After normalization to 18S rRNA, the data are presented with the mean value of the control group set at 100% ( A ). The protein expression of AQP3 in the skin was analyzed by Western blotting. After normalization to glyceraldehyde-3-phosphate dehydrogenase (GAPDH), the data are shown with the mean value of the control group set at 100% (mean ± SD, n = 5; * p

    Journal: Biomolecules

    Article Title: Epidermal Growth Factor Receptor Tyrosine Kinase Inhibitor Erlotinib Induces Dry Skin via Decreased in Aquaporin-3 Expression

    doi: 10.3390/biom10040545

    Figure Lengend Snippet: Expression levels and distribution of aquaporin (AQP) in mouse skin. Erlotinib was administered orally to the mice for 14 days, and the AQP (AQP0-9) mRNA expression level in the skin was measured by real-time RT-PCR. After normalization to 18S rRNA, the data are presented with the mean value of the control group set at 100% ( A ). The protein expression of AQP3 in the skin was analyzed by Western blotting. After normalization to glyceraldehyde-3-phosphate dehydrogenase (GAPDH), the data are shown with the mean value of the control group set at 100% (mean ± SD, n = 5; * p

    Article Snippet: The rabbit anti-rat aquaporin-3 (AQP3) antibody was purchased from Alomone Labs (Jerusalem, Israel).

    Techniques: Expressing, Mouse Assay, Quantitative RT-PCR, Western Blot

    Effect of erlotinib on AQP3 expression in HaCaT cells. Erlotinib was added to HaCaT cells, and the cells were incubated for 24 h. The AQP3 mRNA expression level was measured by real-time RT-PCR. After normalization to RPL30, the data are presented with the mean value of the control group set at 100% ( A ). The protein expression of AQP3 in HaCaT cells was analyzed by Western blotting. After normalization to GAPDH, the data are shown with the mean value of the control group set at 100% ( B ). AQP3 (green) and nuclei (blue) in HaCaT cells were immunostained ( C ). Cell viability was analyzed by the reagent water-soluble tetrazolium salt (WST-1) assay ( D ) (mean ± SD, n = 5; * p

    Journal: Biomolecules

    Article Title: Epidermal Growth Factor Receptor Tyrosine Kinase Inhibitor Erlotinib Induces Dry Skin via Decreased in Aquaporin-3 Expression

    doi: 10.3390/biom10040545

    Figure Lengend Snippet: Effect of erlotinib on AQP3 expression in HaCaT cells. Erlotinib was added to HaCaT cells, and the cells were incubated for 24 h. The AQP3 mRNA expression level was measured by real-time RT-PCR. After normalization to RPL30, the data are presented with the mean value of the control group set at 100% ( A ). The protein expression of AQP3 in HaCaT cells was analyzed by Western blotting. After normalization to GAPDH, the data are shown with the mean value of the control group set at 100% ( B ). AQP3 (green) and nuclei (blue) in HaCaT cells were immunostained ( C ). Cell viability was analyzed by the reagent water-soluble tetrazolium salt (WST-1) assay ( D ) (mean ± SD, n = 5; * p

    Article Snippet: The rabbit anti-rat aquaporin-3 (AQP3) antibody was purchased from Alomone Labs (Jerusalem, Israel).

    Techniques: Expressing, Incubation, Quantitative RT-PCR, Western Blot, WST-1 Assay

    Effect of erlotinib on the phosphorylation of epidermal growth factor receptors (EGFRs) and extracellular signal-regulated kinase (ERK) in HaCaT cells. Erlotinib was added to HaCaT cells, and the cells were incubated for 24 h. The protein expression of phospho (p)-EGFR and p-ERK in HaCaT cells was analyzed by Western blotting. After normalization to EGFR and ERK, the data are shown with the mean value of the control group set at 100% ( A ). Gefitinib was added to HaCaT cells, and the cells were incubated for 24 h. The AQP3 mRNA expression level was measured by real-time RT-PCR. After normalization to RPL30, the data are presented with the mean value of the control group set at 100% ( B ) (mean ± SD, n = 5; * p

    Journal: Biomolecules

    Article Title: Epidermal Growth Factor Receptor Tyrosine Kinase Inhibitor Erlotinib Induces Dry Skin via Decreased in Aquaporin-3 Expression

    doi: 10.3390/biom10040545

    Figure Lengend Snippet: Effect of erlotinib on the phosphorylation of epidermal growth factor receptors (EGFRs) and extracellular signal-regulated kinase (ERK) in HaCaT cells. Erlotinib was added to HaCaT cells, and the cells were incubated for 24 h. The protein expression of phospho (p)-EGFR and p-ERK in HaCaT cells was analyzed by Western blotting. After normalization to EGFR and ERK, the data are shown with the mean value of the control group set at 100% ( A ). Gefitinib was added to HaCaT cells, and the cells were incubated for 24 h. The AQP3 mRNA expression level was measured by real-time RT-PCR. After normalization to RPL30, the data are presented with the mean value of the control group set at 100% ( B ) (mean ± SD, n = 5; * p

    Article Snippet: The rabbit anti-rat aquaporin-3 (AQP3) antibody was purchased from Alomone Labs (Jerusalem, Israel).

    Techniques: Incubation, Expressing, Western Blot, Quantitative RT-PCR