sodium potassium atpase Search Results


antibodies against na k atpase  (Novus Biologicals)
93
Novus Biologicals antibodies against na k atpase
Antibodies Against Na K Atpase, supplied by Novus Biologicals, 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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na k atpase β2 polyclonal  (Alomone Labs)
91
Alomone Labs na k atpase β2 polyclonal
Na K Atpase β2 Polyclonal, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 91/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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v atpase  (Proteintech)
96
Proteintech v atpase
Fig. 6. PPZ treatment induces an increase in lysosomal pH and inhibits the activity of lysosomal enzyme acid phosphatase in KB/V cells. KB/V cells were treated with various concentrations of PPZ (a) or <t>V-ATPase</t> inhibitor Baf-A1 (b) for 4–24 h, and then the supernatant was harvested for pHe measurements. c Effect of PPZ on lysosomal pH in KB/V cells was evaluated by LysoSensor Green probe using fluorescence microscope. d The activity of lysosomal enzyme acid phosphatase in KB/V cells treated with various concentrations of PPZ for 24 h. e Effect of PPZ on the expression of V-ATPase in KB/V cells was detected and quantified using Western blotting analysis. For a, b, d and e, data are presented as the mean ± SD of three separate experiments. *P < 0.05, **P < 0.01 vs. KB/V control group. f Effect of V-ATPase inhibitor Baf-A1 on the synergistic cytotoxicity of PPZ and VCR in KB/V cells. KB/V cells were treated with Baf-A1 (50, 200 nM) with or without PPZ and VCR, then the cell viability was detected by MTT assay. Data are presented as the mean ± SD from three separate experiments. *P < 0.05, **P < 0.01 vs. KB/V control group; △△P < 0.01 vs. PPZ alone group; ##P < 0.01 vs. PPZ and VCR combination group (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article).
V Atpase, supplied by Proteintech, used in various techniques. Bioz Stars score: 96/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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nb300  (Novus Biologicals)
94
Novus Biologicals nb300
Fig. 6. PPZ treatment induces an increase in lysosomal pH and inhibits the activity of lysosomal enzyme acid phosphatase in KB/V cells. KB/V cells were treated with various concentrations of PPZ (a) or <t>V-ATPase</t> inhibitor Baf-A1 (b) for 4–24 h, and then the supernatant was harvested for pHe measurements. c Effect of PPZ on lysosomal pH in KB/V cells was evaluated by LysoSensor Green probe using fluorescence microscope. d The activity of lysosomal enzyme acid phosphatase in KB/V cells treated with various concentrations of PPZ for 24 h. e Effect of PPZ on the expression of V-ATPase in KB/V cells was detected and quantified using Western blotting analysis. For a, b, d and e, data are presented as the mean ± SD of three separate experiments. *P < 0.05, **P < 0.01 vs. KB/V control group. f Effect of V-ATPase inhibitor Baf-A1 on the synergistic cytotoxicity of PPZ and VCR in KB/V cells. KB/V cells were treated with Baf-A1 (50, 200 nM) with or without PPZ and VCR, then the cell viability was detected by MTT assay. Data are presented as the mean ± SD from three separate experiments. *P < 0.05, **P < 0.01 vs. KB/V control group; △△P < 0.01 vs. PPZ alone group; ##P < 0.01 vs. PPZ and VCR combination group (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article).
Nb300, supplied by Novus Biologicals, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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anti na  (Alomone Labs)
93
Alomone Labs anti na
Fig. 6. PPZ treatment induces an increase in lysosomal pH and inhibits the activity of lysosomal enzyme acid phosphatase in KB/V cells. KB/V cells were treated with various concentrations of PPZ (a) or <t>V-ATPase</t> inhibitor Baf-A1 (b) for 4–24 h, and then the supernatant was harvested for pHe measurements. c Effect of PPZ on lysosomal pH in KB/V cells was evaluated by LysoSensor Green probe using fluorescence microscope. d The activity of lysosomal enzyme acid phosphatase in KB/V cells treated with various concentrations of PPZ for 24 h. e Effect of PPZ on the expression of V-ATPase in KB/V cells was detected and quantified using Western blotting analysis. For a, b, d and e, data are presented as the mean ± SD of three separate experiments. *P < 0.05, **P < 0.01 vs. KB/V control group. f Effect of V-ATPase inhibitor Baf-A1 on the synergistic cytotoxicity of PPZ and VCR in KB/V cells. KB/V cells were treated with Baf-A1 (50, 200 nM) with or without PPZ and VCR, then the cell viability was detected by MTT assay. Data are presented as the mean ± SD from three separate experiments. *P < 0.05, **P < 0.01 vs. KB/V control group; △△P < 0.01 vs. PPZ alone group; ##P < 0.01 vs. PPZ and VCR combination group (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article).
Anti Na, 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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na k atpase  (OriGene)
90
OriGene na k atpase
Fig. 6. PPZ treatment induces an increase in lysosomal pH and inhibits the activity of lysosomal enzyme acid phosphatase in KB/V cells. KB/V cells were treated with various concentrations of PPZ (a) or <t>V-ATPase</t> inhibitor Baf-A1 (b) for 4–24 h, and then the supernatant was harvested for pHe measurements. c Effect of PPZ on lysosomal pH in KB/V cells was evaluated by LysoSensor Green probe using fluorescence microscope. d The activity of lysosomal enzyme acid phosphatase in KB/V cells treated with various concentrations of PPZ for 24 h. e Effect of PPZ on the expression of V-ATPase in KB/V cells was detected and quantified using Western blotting analysis. For a, b, d and e, data are presented as the mean ± SD of three separate experiments. *P < 0.05, **P < 0.01 vs. KB/V control group. f Effect of V-ATPase inhibitor Baf-A1 on the synergistic cytotoxicity of PPZ and VCR in KB/V cells. KB/V cells were treated with Baf-A1 (50, 200 nM) with or without PPZ and VCR, then the cell viability was detected by MTT assay. Data are presented as the mean ± SD from three separate experiments. *P < 0.05, **P < 0.01 vs. KB/V control group; △△P < 0.01 vs. PPZ alone group; ##P < 0.01 vs. PPZ and VCR combination group (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article).
Na K Atpase, supplied by OriGene, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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anti atp1a3 mouse nb300540 novus biologicals  (Novus Biologicals)
93
Novus Biologicals anti atp1a3 mouse nb300540 novus biologicals
Fig. 6. PPZ treatment induces an increase in lysosomal pH and inhibits the activity of lysosomal enzyme acid phosphatase in KB/V cells. KB/V cells were treated with various concentrations of PPZ (a) or <t>V-ATPase</t> inhibitor Baf-A1 (b) for 4–24 h, and then the supernatant was harvested for pHe measurements. c Effect of PPZ on lysosomal pH in KB/V cells was evaluated by LysoSensor Green probe using fluorescence microscope. d The activity of lysosomal enzyme acid phosphatase in KB/V cells treated with various concentrations of PPZ for 24 h. e Effect of PPZ on the expression of V-ATPase in KB/V cells was detected and quantified using Western blotting analysis. For a, b, d and e, data are presented as the mean ± SD of three separate experiments. *P < 0.05, **P < 0.01 vs. KB/V control group. f Effect of V-ATPase inhibitor Baf-A1 on the synergistic cytotoxicity of PPZ and VCR in KB/V cells. KB/V cells were treated with Baf-A1 (50, 200 nM) with or without PPZ and VCR, then the cell viability was detected by MTT assay. Data are presented as the mean ± SD from three separate experiments. *P < 0.05, **P < 0.01 vs. KB/V control group; △△P < 0.01 vs. PPZ alone group; ##P < 0.01 vs. PPZ and VCR combination group (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article).
Anti Atp1a3 Mouse Nb300540 Novus Biologicals, supplied by Novus Biologicals, 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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Average 93 stars, based on 1 article reviews
anti atp1a3 mouse nb300540 novus biologicals - by Bioz Stars, 2026-03
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na k atpase α subunit  (Cusabio)
92
Cusabio na k atpase α subunit
Fig. 6. PPZ treatment induces an increase in lysosomal pH and inhibits the activity of lysosomal enzyme acid phosphatase in KB/V cells. KB/V cells were treated with various concentrations of PPZ (a) or <t>V-ATPase</t> inhibitor Baf-A1 (b) for 4–24 h, and then the supernatant was harvested for pHe measurements. c Effect of PPZ on lysosomal pH in KB/V cells was evaluated by LysoSensor Green probe using fluorescence microscope. d The activity of lysosomal enzyme acid phosphatase in KB/V cells treated with various concentrations of PPZ for 24 h. e Effect of PPZ on the expression of V-ATPase in KB/V cells was detected and quantified using Western blotting analysis. For a, b, d and e, data are presented as the mean ± SD of three separate experiments. *P < 0.05, **P < 0.01 vs. KB/V control group. f Effect of V-ATPase inhibitor Baf-A1 on the synergistic cytotoxicity of PPZ and VCR in KB/V cells. KB/V cells were treated with Baf-A1 (50, 200 nM) with or without PPZ and VCR, then the cell viability was detected by MTT assay. Data are presented as the mean ± SD from three separate experiments. *P < 0.05, **P < 0.01 vs. KB/V control group; △△P < 0.01 vs. PPZ alone group; ##P < 0.01 vs. PPZ and VCR combination group (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article).
Na K Atpase α Subunit, supplied by Cusabio, 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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anti na k atpase  (Alomone Labs)
91
Alomone Labs anti na k atpase
Fig. 6. PPZ treatment induces an increase in lysosomal pH and inhibits the activity of lysosomal enzyme acid phosphatase in KB/V cells. KB/V cells were treated with various concentrations of PPZ (a) or <t>V-ATPase</t> inhibitor Baf-A1 (b) for 4–24 h, and then the supernatant was harvested for pHe measurements. c Effect of PPZ on lysosomal pH in KB/V cells was evaluated by LysoSensor Green probe using fluorescence microscope. d The activity of lysosomal enzyme acid phosphatase in KB/V cells treated with various concentrations of PPZ for 24 h. e Effect of PPZ on the expression of V-ATPase in KB/V cells was detected and quantified using Western blotting analysis. For a, b, d and e, data are presented as the mean ± SD of three separate experiments. *P < 0.05, **P < 0.01 vs. KB/V control group. f Effect of V-ATPase inhibitor Baf-A1 on the synergistic cytotoxicity of PPZ and VCR in KB/V cells. KB/V cells were treated with Baf-A1 (50, 200 nM) with or without PPZ and VCR, then the cell viability was detected by MTT assay. Data are presented as the mean ± SD from three separate experiments. *P < 0.05, **P < 0.01 vs. KB/V control group; △△P < 0.01 vs. PPZ alone group; ##P < 0.01 vs. PPZ and VCR combination group (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article).
Anti Na K Atpase, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 91/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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rabbit anti na k atpase alpha1 subunit  (Novus Biologicals)
90
Novus Biologicals rabbit anti na k atpase alpha1 subunit
Fig. 6. PPZ treatment induces an increase in lysosomal pH and inhibits the activity of lysosomal enzyme acid phosphatase in KB/V cells. KB/V cells were treated with various concentrations of PPZ (a) or <t>V-ATPase</t> inhibitor Baf-A1 (b) for 4–24 h, and then the supernatant was harvested for pHe measurements. c Effect of PPZ on lysosomal pH in KB/V cells was evaluated by LysoSensor Green probe using fluorescence microscope. d The activity of lysosomal enzyme acid phosphatase in KB/V cells treated with various concentrations of PPZ for 24 h. e Effect of PPZ on the expression of V-ATPase in KB/V cells was detected and quantified using Western blotting analysis. For a, b, d and e, data are presented as the mean ± SD of three separate experiments. *P < 0.05, **P < 0.01 vs. KB/V control group. f Effect of V-ATPase inhibitor Baf-A1 on the synergistic cytotoxicity of PPZ and VCR in KB/V cells. KB/V cells were treated with Baf-A1 (50, 200 nM) with or without PPZ and VCR, then the cell viability was detected by MTT assay. Data are presented as the mean ± SD from three separate experiments. *P < 0.05, **P < 0.01 vs. KB/V control group; △△P < 0.01 vs. PPZ alone group; ##P < 0.01 vs. PPZ and VCR combination group (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article).
Rabbit Anti Na K Atpase Alpha1 Subunit, supplied by Novus Biologicals, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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anti atp1a3 antibody  (Novus Biologicals)
92
Novus Biologicals anti atp1a3 antibody
Fig. 1. <t>ATP1A3</t> is a neuron-specific EV marker candidate besides NCAM1 and L1CAM and associated with EVs in human brain tissues and biofluids at a single- particle level. (A) Experimental design for evaluating whether NDEV marker candidates (ATP1A3, NCAM1, and L1CAM) is associated with EVs. UC, ultracentrifugation; SU- GC, sucrose gradient centrifugation; SEC, size exclusion chromatography. (B) Transmission electron micrograph showing immunogold labeling of EVs isolated from iNeuron, brain, CSF, and plasma stained with anti-CD9 antibody (10 nm) and anti-ATP1A3, NCAM1, and L1CAM antibody (5 nm). Scale bars, 100 nm. (C) Representative fluorescent images of iNeuron-EV detected by fluorescent-conjugated antibodies (red: ATP1A3-AF647, NCAM1-AF647 or L1CAM-AF647; blue: CD9/81C/63-AF488) on Exoview chip CD63 capture spots. The round circle in magnified images masks costained particles with tetraspanins. Scale bars, 25 and 5 μm. (D) Percentages of ATP1A3, NCAM1, and L1CAM fluorescent particle counts relative to total particles in iNeuron derived EVs (n = 2 biological replicates) on Exoview chip tetraspanins capture spots. (E) Representative fluorescent images of brain- and CSF-EV detected by fluorescent-conjugated antibodies (green: ATP1A3-AF555; red: NCAM1-AF647 or L1CAM-AF647; blue: CD9/81/63-AF488) on Exoview chip CD9 capture spots. The round circle in magnified images masks co-stained particles with tetraspanins. Scale bars, 25 and 5 μm. (F) Percentages of ATP1A3, NCAM1, and L1CAM fluorescent particle counts relative to total particles in brain (n = 2) and CSF (n = 2) derived EVs on Exoview chip tetraspanins capture spots.
Anti Atp1a3 Antibody, supplied by Novus Biologicals, 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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Image Search Results


Fig. 6. PPZ treatment induces an increase in lysosomal pH and inhibits the activity of lysosomal enzyme acid phosphatase in KB/V cells. KB/V cells were treated with various concentrations of PPZ (a) or V-ATPase inhibitor Baf-A1 (b) for 4–24 h, and then the supernatant was harvested for pHe measurements. c Effect of PPZ on lysosomal pH in KB/V cells was evaluated by LysoSensor Green probe using fluorescence microscope. d The activity of lysosomal enzyme acid phosphatase in KB/V cells treated with various concentrations of PPZ for 24 h. e Effect of PPZ on the expression of V-ATPase in KB/V cells was detected and quantified using Western blotting analysis. For a, b, d and e, data are presented as the mean ± SD of three separate experiments. *P < 0.05, **P < 0.01 vs. KB/V control group. f Effect of V-ATPase inhibitor Baf-A1 on the synergistic cytotoxicity of PPZ and VCR in KB/V cells. KB/V cells were treated with Baf-A1 (50, 200 nM) with or without PPZ and VCR, then the cell viability was detected by MTT assay. Data are presented as the mean ± SD from three separate experiments. *P < 0.05, **P < 0.01 vs. KB/V control group; △△P < 0.01 vs. PPZ alone group; ##P < 0.01 vs. PPZ and VCR combination group (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article).

Journal: Biomedicine & pharmacotherapy = Biomedecine & pharmacotherapie

Article Title: Pantoprazole pretreatment elevates sensitivity to vincristine in drug-resistant oral epidermoid carcinoma in vitro and in vivo.

doi: 10.1016/j.biopha.2019.109478

Figure Lengend Snippet: Fig. 6. PPZ treatment induces an increase in lysosomal pH and inhibits the activity of lysosomal enzyme acid phosphatase in KB/V cells. KB/V cells were treated with various concentrations of PPZ (a) or V-ATPase inhibitor Baf-A1 (b) for 4–24 h, and then the supernatant was harvested for pHe measurements. c Effect of PPZ on lysosomal pH in KB/V cells was evaluated by LysoSensor Green probe using fluorescence microscope. d The activity of lysosomal enzyme acid phosphatase in KB/V cells treated with various concentrations of PPZ for 24 h. e Effect of PPZ on the expression of V-ATPase in KB/V cells was detected and quantified using Western blotting analysis. For a, b, d and e, data are presented as the mean ± SD of three separate experiments. *P < 0.05, **P < 0.01 vs. KB/V control group. f Effect of V-ATPase inhibitor Baf-A1 on the synergistic cytotoxicity of PPZ and VCR in KB/V cells. KB/V cells were treated with Baf-A1 (50, 200 nM) with or without PPZ and VCR, then the cell viability was detected by MTT assay. Data are presented as the mean ± SD from three separate experiments. *P < 0.05, **P < 0.01 vs. KB/V control group; △△P < 0.01 vs. PPZ alone group; ##P < 0.01 vs. PPZ and VCR combination group (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article).

Article Snippet: Membranes were blocked in 5% (w/v) non-fat milk before incubation with primary antibodies (1:1000 in primary antibody diluent) against PARP (#9532), cleavedcaspase 3 (#9664), Bcl-xL(#2762), Bcl-2 (#2872), Bax (#2772), p21 (#2947), Cyclin B1 (#4138), cdc2 (#9112), p-cdc2 (#9111), MMP2 (#4022), MMP9 (#3852), P-gp (#13342), EGFR (#2232), p-EGFR (#2235), p-ERK (#4370), p-p38 (#4511), PI3K (#3358), Akt (#2965), p-Akt (#2965), mTOR (#2972), p-mTOR (#2971), β-actin (#4970) (all from Cell Signaling Technology, CST, Boston, MA) and V-ATPase (14418-1-AP, Proteintech, Wuhan, China) at 4 °C overnight.

Techniques: Activity Assay, Microscopy, Expressing, Western Blot, Control, MTT Assay

Fig. 1. ATP1A3 is a neuron-specific EV marker candidate besides NCAM1 and L1CAM and associated with EVs in human brain tissues and biofluids at a single- particle level. (A) Experimental design for evaluating whether NDEV marker candidates (ATP1A3, NCAM1, and L1CAM) is associated with EVs. UC, ultracentrifugation; SU- GC, sucrose gradient centrifugation; SEC, size exclusion chromatography. (B) Transmission electron micrograph showing immunogold labeling of EVs isolated from iNeuron, brain, CSF, and plasma stained with anti-CD9 antibody (10 nm) and anti-ATP1A3, NCAM1, and L1CAM antibody (5 nm). Scale bars, 100 nm. (C) Representative fluorescent images of iNeuron-EV detected by fluorescent-conjugated antibodies (red: ATP1A3-AF647, NCAM1-AF647 or L1CAM-AF647; blue: CD9/81C/63-AF488) on Exoview chip CD63 capture spots. The round circle in magnified images masks costained particles with tetraspanins. Scale bars, 25 and 5 μm. (D) Percentages of ATP1A3, NCAM1, and L1CAM fluorescent particle counts relative to total particles in iNeuron derived EVs (n = 2 biological replicates) on Exoview chip tetraspanins capture spots. (E) Representative fluorescent images of brain- and CSF-EV detected by fluorescent-conjugated antibodies (green: ATP1A3-AF555; red: NCAM1-AF647 or L1CAM-AF647; blue: CD9/81/63-AF488) on Exoview chip CD9 capture spots. The round circle in magnified images masks co-stained particles with tetraspanins. Scale bars, 25 and 5 μm. (F) Percentages of ATP1A3, NCAM1, and L1CAM fluorescent particle counts relative to total particles in brain (n = 2) and CSF (n = 2) derived EVs on Exoview chip tetraspanins capture spots.

Journal: Science advances

Article Title: ATP1A3 as a target for isolating neuron-specific extracellular vesicles from human brain and biofluids.

doi: 10.1126/sciadv.adi3647

Figure Lengend Snippet: Fig. 1. ATP1A3 is a neuron-specific EV marker candidate besides NCAM1 and L1CAM and associated with EVs in human brain tissues and biofluids at a single- particle level. (A) Experimental design for evaluating whether NDEV marker candidates (ATP1A3, NCAM1, and L1CAM) is associated with EVs. UC, ultracentrifugation; SU- GC, sucrose gradient centrifugation; SEC, size exclusion chromatography. (B) Transmission electron micrograph showing immunogold labeling of EVs isolated from iNeuron, brain, CSF, and plasma stained with anti-CD9 antibody (10 nm) and anti-ATP1A3, NCAM1, and L1CAM antibody (5 nm). Scale bars, 100 nm. (C) Representative fluorescent images of iNeuron-EV detected by fluorescent-conjugated antibodies (red: ATP1A3-AF647, NCAM1-AF647 or L1CAM-AF647; blue: CD9/81C/63-AF488) on Exoview chip CD63 capture spots. The round circle in magnified images masks costained particles with tetraspanins. Scale bars, 25 and 5 μm. (D) Percentages of ATP1A3, NCAM1, and L1CAM fluorescent particle counts relative to total particles in iNeuron derived EVs (n = 2 biological replicates) on Exoview chip tetraspanins capture spots. (E) Representative fluorescent images of brain- and CSF-EV detected by fluorescent-conjugated antibodies (green: ATP1A3-AF555; red: NCAM1-AF647 or L1CAM-AF647; blue: CD9/81/63-AF488) on Exoview chip CD9 capture spots. The round circle in magnified images masks co-stained particles with tetraspanins. Scale bars, 25 and 5 μm. (F) Percentages of ATP1A3, NCAM1, and L1CAM fluorescent particle counts relative to total particles in brain (n = 2) and CSF (n = 2) derived EVs on Exoview chip tetraspanins capture spots.

Article Snippet: In our experiment, EV antibodies cocktail (EV-TC-AB-01, NanoView Biosciences) containing AF488-conjugated anti-CD63, anti-CD81, and anti-CD9 antibodies, AF647-conjugated anti-ATP1A3 antibody (NBP299238AF647, Novus Biologicals), AF647-conjugated anti-NCAM1 antibody (catalog no. 563443, BD Biosciences), and AF647-conjugated anti-L1CAM antibody (catalog no. 564194, BD Biosciences) were used for detection of EV subpopulations.

Techniques: Marker, Single Particle, Gradient Centrifugation, Size-exclusion Chromatography, Transmission Assay, Labeling, Isolation, Clinical Proteomics, Staining, Derivative Assay

Fig. 2. Comparative analysis of NDEV candidate markers (ATP1A3, NCAM1, and L1CAM) expression in EVs by super-resolution microscopy. (A) Schematics showing the staining protocol of ATP1A3 and NCAM1 for EVs. (B) Representative super-resolution microscopy images showing the expression of ATP1A3 and NCAM1 in pan-tetraspanins (CD9/81/63) positive EVs at a single-particle level. Scale bars, 100 nm. (C) Pie charts showing the heterogeneous EV population with ATP1A3 and NCAM1 in different EV preparations. Particles with diameter less than 200 nm were quantified. (D) Schematics showing the staining protocol of ATP1A3 and L1CAM for EVs. (E) Representative super-resolution microscopy images showing the expression of ATP1A3 and L1CAM in pan-tetraspanins (CD9/81/63) positive EVs at a single- particle level. Scale bars, 100 nm. (F) Pie charts showing the heterogeneous EV population with ATP1A3 and L1CAM in different EV preparations. Particles with diameter less than 200 nm were quantified.

Journal: Science advances

Article Title: ATP1A3 as a target for isolating neuron-specific extracellular vesicles from human brain and biofluids.

doi: 10.1126/sciadv.adi3647

Figure Lengend Snippet: Fig. 2. Comparative analysis of NDEV candidate markers (ATP1A3, NCAM1, and L1CAM) expression in EVs by super-resolution microscopy. (A) Schematics showing the staining protocol of ATP1A3 and NCAM1 for EVs. (B) Representative super-resolution microscopy images showing the expression of ATP1A3 and NCAM1 in pan-tetraspanins (CD9/81/63) positive EVs at a single-particle level. Scale bars, 100 nm. (C) Pie charts showing the heterogeneous EV population with ATP1A3 and NCAM1 in different EV preparations. Particles with diameter less than 200 nm were quantified. (D) Schematics showing the staining protocol of ATP1A3 and L1CAM for EVs. (E) Representative super-resolution microscopy images showing the expression of ATP1A3 and L1CAM in pan-tetraspanins (CD9/81/63) positive EVs at a single- particle level. Scale bars, 100 nm. (F) Pie charts showing the heterogeneous EV population with ATP1A3 and L1CAM in different EV preparations. Particles with diameter less than 200 nm were quantified.

Article Snippet: In our experiment, EV antibodies cocktail (EV-TC-AB-01, NanoView Biosciences) containing AF488-conjugated anti-CD63, anti-CD81, and anti-CD9 antibodies, AF647-conjugated anti-ATP1A3 antibody (NBP299238AF647, Novus Biologicals), AF647-conjugated anti-NCAM1 antibody (catalog no. 563443, BD Biosciences), and AF647-conjugated anti-L1CAM antibody (catalog no. 564194, BD Biosciences) were used for detection of EV subpopulations.

Techniques: Expressing, Super-Resolution Microscopy, Staining, Single Particle

Fig. 3. ATP1A3+ EV immunoprecipitated from human brain tissues displays higher neuronal cell type specificity. (A) Workflow for characterizing ATP1A3+, NCAM1+, and L1CAM+ EVs immunoprecipitated from brain. (B) Western blot images of 5% total EVs (Input), supernatant (S.P), and immunoprecipitated EVs (A, ATP1A3; L, L1CAM; N, NCAM1) isolated from control brains (n = 3) and probed for neuron and glia markers. Equivalent proteins from brain-EVs were used for immuno- capturing ATP1A3, L1CAM, and NCAM1. IB, immunoblotting antibody (C) Principal components analysis (PCA). The EV proteome of total brain-EV and immunoprecip- itated EVs were measured in triplicate and classified. (D) Venn diagram showing the number of proteins differentially identified in total, ATP1A3+, L1CAM+, and NCAM1+

Journal: Science advances

Article Title: ATP1A3 as a target for isolating neuron-specific extracellular vesicles from human brain and biofluids.

doi: 10.1126/sciadv.adi3647

Figure Lengend Snippet: Fig. 3. ATP1A3+ EV immunoprecipitated from human brain tissues displays higher neuronal cell type specificity. (A) Workflow for characterizing ATP1A3+, NCAM1+, and L1CAM+ EVs immunoprecipitated from brain. (B) Western blot images of 5% total EVs (Input), supernatant (S.P), and immunoprecipitated EVs (A, ATP1A3; L, L1CAM; N, NCAM1) isolated from control brains (n = 3) and probed for neuron and glia markers. Equivalent proteins from brain-EVs were used for immuno- capturing ATP1A3, L1CAM, and NCAM1. IB, immunoblotting antibody (C) Principal components analysis (PCA). The EV proteome of total brain-EV and immunoprecip- itated EVs were measured in triplicate and classified. (D) Venn diagram showing the number of proteins differentially identified in total, ATP1A3+, L1CAM+, and NCAM1+

Article Snippet: In our experiment, EV antibodies cocktail (EV-TC-AB-01, NanoView Biosciences) containing AF488-conjugated anti-CD63, anti-CD81, and anti-CD9 antibodies, AF647-conjugated anti-ATP1A3 antibody (NBP299238AF647, Novus Biologicals), AF647-conjugated anti-NCAM1 antibody (catalog no. 563443, BD Biosciences), and AF647-conjugated anti-L1CAM antibody (catalog no. 564194, BD Biosciences) were used for detection of EV subpopulations.

Techniques: Immunoprecipitation, Western Blot, Isolation, Control

Fig. 4. ATP1A3+ EV analysis in human CSF and plasma. (A) Workflow for characterizing ATP1A3+ EVs immunoprecipitated from CSF. (B) Simoa quantification of AD related neuropathogenic proteins (Aβ42, Aβ40, T-tau, P-tau181, and SNAP25) in ATP1A3+ CSF-EVs from healthy controls (n = 3) and AD (n = 3). Data are presented by box and whisker plot which shows the median and the 25th to 75th percentiles. (C) Aβ42/Aβ40 ratio in CSF, total CSF-EV, and ATP1A3 immunoprecipitation (IP) CSF-EV samples among AD group. (D) Workflow explaining single-particle analysis of Aβ+ population in ATP1A3+ plasma-EV to distinguish AD from healthy controls (CTL) and mild cognitive impairment (MCI). (E) Representative super-resolution microscopy images showing plasma-derived Aβ+ ATP1A3+ single EV among control, MCI, and AD groups [green: FITC-CD9/81/63, red: Alexa Flour 555-ATP1A3, gray: Alexa Flour 647-Aβ (clone 4G8)]. Scale bars, 100 nm. (F) Percentage of Aβ+ population in ATP1A3+ plasma-EV among control, MCI, and AD groups (n = 10 per group). (G) Correlations of Aβ+ population in ATP1A3+ plasma-EV with clinical Braak stage, Mini- Mental State Examination (MMSE), and Dementia Rating Scale (DRS) scores in combined control (n = 7), MCI (n = 10), and AD (n = 10) samples. The dashed line indicates 95% confidence band of the best-fit line. Pearson correlation coefficients are shown. (H) Quantification of Aβ42/Aβ40 level in total plasma samples from CTL (n = 9), MCI (n = 10), and AD (n = 10). (I to J) Receiver-operating characteristics (ROC) curves displaying the performance of Aβ+ population in ATP1A3+ plasma-EV, plasma Aβ40, Aβ42, Aβ42/Aβ40 ratio, and T-tau to distinguish (I) AD from CTL and (J) AD from MCI. T-tau, total tau; AUC, area under the curve; CI, confidence interval; ns, no significance; *P < 0.05, **P < 0.01, ***P < 0.001.

Journal: Science advances

Article Title: ATP1A3 as a target for isolating neuron-specific extracellular vesicles from human brain and biofluids.

doi: 10.1126/sciadv.adi3647

Figure Lengend Snippet: Fig. 4. ATP1A3+ EV analysis in human CSF and plasma. (A) Workflow for characterizing ATP1A3+ EVs immunoprecipitated from CSF. (B) Simoa quantification of AD related neuropathogenic proteins (Aβ42, Aβ40, T-tau, P-tau181, and SNAP25) in ATP1A3+ CSF-EVs from healthy controls (n = 3) and AD (n = 3). Data are presented by box and whisker plot which shows the median and the 25th to 75th percentiles. (C) Aβ42/Aβ40 ratio in CSF, total CSF-EV, and ATP1A3 immunoprecipitation (IP) CSF-EV samples among AD group. (D) Workflow explaining single-particle analysis of Aβ+ population in ATP1A3+ plasma-EV to distinguish AD from healthy controls (CTL) and mild cognitive impairment (MCI). (E) Representative super-resolution microscopy images showing plasma-derived Aβ+ ATP1A3+ single EV among control, MCI, and AD groups [green: FITC-CD9/81/63, red: Alexa Flour 555-ATP1A3, gray: Alexa Flour 647-Aβ (clone 4G8)]. Scale bars, 100 nm. (F) Percentage of Aβ+ population in ATP1A3+ plasma-EV among control, MCI, and AD groups (n = 10 per group). (G) Correlations of Aβ+ population in ATP1A3+ plasma-EV with clinical Braak stage, Mini- Mental State Examination (MMSE), and Dementia Rating Scale (DRS) scores in combined control (n = 7), MCI (n = 10), and AD (n = 10) samples. The dashed line indicates 95% confidence band of the best-fit line. Pearson correlation coefficients are shown. (H) Quantification of Aβ42/Aβ40 level in total plasma samples from CTL (n = 9), MCI (n = 10), and AD (n = 10). (I to J) Receiver-operating characteristics (ROC) curves displaying the performance of Aβ+ population in ATP1A3+ plasma-EV, plasma Aβ40, Aβ42, Aβ42/Aβ40 ratio, and T-tau to distinguish (I) AD from CTL and (J) AD from MCI. T-tau, total tau; AUC, area under the curve; CI, confidence interval; ns, no significance; *P < 0.05, **P < 0.01, ***P < 0.001.

Article Snippet: In our experiment, EV antibodies cocktail (EV-TC-AB-01, NanoView Biosciences) containing AF488-conjugated anti-CD63, anti-CD81, and anti-CD9 antibodies, AF647-conjugated anti-ATP1A3 antibody (NBP299238AF647, Novus Biologicals), AF647-conjugated anti-NCAM1 antibody (catalog no. 563443, BD Biosciences), and AF647-conjugated anti-L1CAM antibody (catalog no. 564194, BD Biosciences) were used for detection of EV subpopulations.

Techniques: Clinical Proteomics, Immunoprecipitation, Whisker Assay, Single Particle, Super-Resolution Microscopy, Derivative Assay, Control