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encodes aav8 capsid protein  (Addgene inc)


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    Addgene inc encodes aav8 capsid protein
    Figure 5. BEC-enhancer AAV labels brain endothelial cells in brain blood vessels in vivo (A) Schematic experimental design to visualize endothelial cells in vivo. Mice received retro-orbital injection of BEC-enhancer AAV and <t>AAV8-P3-Alb-mScarlet</t> to label albumin in the blood and visualize brain blood vessels in vivo. The graphics were created with BioRender.com. (B) Representative in vivo two-photon imaging of anesthetized mice 4 weeks after AAV injection. Top left panel with 1-s averaged pixel density from blood albumin signal detected at single plane from somatosensory cortex in vivo. Top right panel with 1-s averaged pixel density from brain endothelial cells labeled by AAV- PHP.eB-BEC enhancer-minBG-SYFP2 from the same plane as the top left panel. Bottom, the merged image from both panels (see Video S2). (C) Representative in vivo two-photon imaging of smaller blood vessels relative to those shown in (B), detected for both BEC-enhancer AAV and AAV8-P3-Alb- mScarlet signals. (D) Representative in vivo two-photon imaging of anesthetized mice 10 weeks after AAV injection. Top panel with 5-s averaged pixel density from blood albumin signal depicted in magenta at a single plane from hippocampal CA1 in vivo. Middle panel with 5-s averaged pixel density from brain endothelial cells labeled by AAV-PHP.eB-BEC enhancer-minBG-SYFP2 (BEC-enhancer AAV, green) from the same plane as the top left panel. Bottom, the merged image of the above panels. The albumin signal as well as the signal from transduced cells with BEC-enhancer AAV is stable for at least a couple of months. Some speckles are due to debris artifacts.
    Encodes Aav8 Capsid Protein, supplied by Addgene inc, used in various techniques. Bioz Stars score: 96/100, based on 140 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    1) Product Images from "Specific targeting of brain endothelial cells using enhancer AAV vectors."

    Article Title: Specific targeting of brain endothelial cells using enhancer AAV vectors.

    Journal: Neuron

    doi: 10.1016/j.neuron.2025.03.031

    Figure 5. BEC-enhancer AAV labels brain endothelial cells in brain blood vessels in vivo (A) Schematic experimental design to visualize endothelial cells in vivo. Mice received retro-orbital injection of BEC-enhancer AAV and AAV8-P3-Alb-mScarlet to label albumin in the blood and visualize brain blood vessels in vivo. The graphics were created with BioRender.com. (B) Representative in vivo two-photon imaging of anesthetized mice 4 weeks after AAV injection. Top left panel with 1-s averaged pixel density from blood albumin signal detected at single plane from somatosensory cortex in vivo. Top right panel with 1-s averaged pixel density from brain endothelial cells labeled by AAV- PHP.eB-BEC enhancer-minBG-SYFP2 from the same plane as the top left panel. Bottom, the merged image from both panels (see Video S2). (C) Representative in vivo two-photon imaging of smaller blood vessels relative to those shown in (B), detected for both BEC-enhancer AAV and AAV8-P3-Alb- mScarlet signals. (D) Representative in vivo two-photon imaging of anesthetized mice 10 weeks after AAV injection. Top panel with 5-s averaged pixel density from blood albumin signal depicted in magenta at a single plane from hippocampal CA1 in vivo. Middle panel with 5-s averaged pixel density from brain endothelial cells labeled by AAV-PHP.eB-BEC enhancer-minBG-SYFP2 (BEC-enhancer AAV, green) from the same plane as the top left panel. Bottom, the merged image of the above panels. The albumin signal as well as the signal from transduced cells with BEC-enhancer AAV is stable for at least a couple of months. Some speckles are due to debris artifacts.
    Figure Legend Snippet: Figure 5. BEC-enhancer AAV labels brain endothelial cells in brain blood vessels in vivo (A) Schematic experimental design to visualize endothelial cells in vivo. Mice received retro-orbital injection of BEC-enhancer AAV and AAV8-P3-Alb-mScarlet to label albumin in the blood and visualize brain blood vessels in vivo. The graphics were created with BioRender.com. (B) Representative in vivo two-photon imaging of anesthetized mice 4 weeks after AAV injection. Top left panel with 1-s averaged pixel density from blood albumin signal detected at single plane from somatosensory cortex in vivo. Top right panel with 1-s averaged pixel density from brain endothelial cells labeled by AAV- PHP.eB-BEC enhancer-minBG-SYFP2 from the same plane as the top left panel. Bottom, the merged image from both panels (see Video S2). (C) Representative in vivo two-photon imaging of smaller blood vessels relative to those shown in (B), detected for both BEC-enhancer AAV and AAV8-P3-Alb- mScarlet signals. (D) Representative in vivo two-photon imaging of anesthetized mice 10 weeks after AAV injection. Top panel with 5-s averaged pixel density from blood albumin signal depicted in magenta at a single plane from hippocampal CA1 in vivo. Middle panel with 5-s averaged pixel density from brain endothelial cells labeled by AAV-PHP.eB-BEC enhancer-minBG-SYFP2 (BEC-enhancer AAV, green) from the same plane as the top left panel. Bottom, the merged image of the above panels. The albumin signal as well as the signal from transduced cells with BEC-enhancer AAV is stable for at least a couple of months. Some speckles are due to debris artifacts.

    Techniques Used: In Vivo, Injection, Imaging, Labeling

    Related Articles

    other:

    Article Title: Specific targeting of brain endothelial cells using enhancer AAV vectors.
    Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER SciPy http://www.scipy.org/ RRID:SCR_008058 Insight Segmentation and Registration Toolkit http://www.itk.org RRID:SCR_001149 scikit-image https://scikit-image.org/ RRID:SCR_021142 Blood vessel processing code This paper https://doi.org/10.5281/zenodo.14876892 GraphPad Prism 10.4.1 http://www.graphpad.com/ RRID:SCR_002798

    Virus:

    Article Title: Specific targeting of brain endothelial cells using enhancer AAV vectors.
    Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Ethidium Bromide (for maxiprep plasmids) Sigma CAS #: 1239-45-8 Isopropanol (for maxiprep plasmids) Fisher Scientific CAS #: 67-63-0 Phenol (for maxiprep plasmids) Fisher Scientific CAS #: 108-95-2 Chloroform (for maxiprep plasmids) Fisher Scientific CAS #: 67-66-3 Critical commercial assays Pierce Silver Stain Kit (AAV purification) Thermo Fisher Cat#: 24612 Deposited data CATlas http://catlas.org/mousebrain/#! .. RRID:SCR_018690 Mendeley Data: quantification and metadata This paper https://doi.org/10.17632/5kvfknrgng.1 UCSC Genome Browser http://genome.ucsc.edu/ RRID:SCR_005780 Experimental models: Cell lines Cells: AAVpro 293T (for virus packaging) Takara Bio RRID:CVCL_B0XW Experimental models: Organisms/strains Mouse: Tie2-Cre (B6.Cg-Tg(Tek-cre)1Ywa/J) The Jackson Laboratory RRID:IMSR_JAX:008863 Mouse: Ai9 (B6.Cg-Gt(ROSA)26Sortm9 (CAG-tdTomato)Hze/J) The Jackson Laboratory RRID:IMSR_JAX:007909 Mouse: 5xFAD (B6.Cg-Tg (APPSwFlLon,PSEN1*M146L*L286V)6799Vas/Mmjax) The Jackson Laboratory RRID:MMRRC_034848-JAX Mouse: B6J (C57BL/6J) The Jackson Laboratory RRID:IMSR_JAX:000664 Oligonucleotides Fwd ITR primer, 50-GGAACCCCTAGTGATGGAGTT Aurnhammer et al.68 PMID: 22428977 Rev ITR primer, 50-CGGCCTCAGTGAGCGA Aurnhammer et al.68 PMID: 22428977 PowerUP SYBR green Master mix Thermo Fisher A46012 Recombinant DNA pUCmini-iCAP-PHP.eB plasmid (encodes engineered PHP.eB capsid protein) Addgene Cat#: 103005; RRID:Addgene_103005 pAAV2/8 (encodes AAV8 capsid protein) Addgene Cat#: 112864; RRID:Addgene_112864 pAAV-BR1 Körbelin et al.13 PMID: 27137490 Ad helper plasmid (mini-pHelper) This paper N/A pAAV-BEC enhancer-minBG-SYFP2 This paper Cat #: 236722; RRID:Addgene_236722 pAAV-minBG-SYFP2 This paper Cat #: 236723; RRID:Addgene_236723 pAAV-CAG-mTurquoise2 Addgene Cat#: 99122; RRID:Addgene_99122 pAAV-BEC enhancer-minBG-iCre This paper Cat #: 236724; RRID:Addgene_236724 pAAV-BEC enhancer-minBG-iCre(R279T)_4x6T This paper Cat #: 236727; RRID:Addgene_236727 P3-Alb-mScarlet Addgene Cat#: 183461; RRID:Addgene_183461 AiP14187 - pAAV-AiE0459m_3xC2-minBGiCre(R297T)-BGHpA (plasmid used to clone out attenuated iCre(R279T)) Addgene Cat#: 229920; RRID:Addgene_229920 AAV-GfaABC1D-ERCreER-4x6T (plasmid used to clone out 4x6T) Addgene Cat#: 196411; RRID:Addgene_196411 Software and algorithms ImageJ (v. 1.53c) https://imagej.net/ RRID:SCR_003070 Python Programming Language http://www.python.org/ RRID:SCR_008394 OpenCV http://opencv.org RRID:SCR_015526 MatPlotLib http://matplotlib.sourceforge.net RRID:SCR_008624 NumPy http://www.numpy.org RRID:SCR_008633 SimpleITK https://simpleitk.org/ RRID:SCR_024693 (Continued on next page) e2 Neuron 113, 1562–1578.e1–e6, May 21, 2025 .. REAGENT or RESOURCE SOURCE IDENTIFIER SciPy http://www.scipy.org/ RRID:SCR_008058 Insight Segmentation and Registration Toolkit http://www.itk.org RRID:SCR_001149 scikit-image https://scikit-image.org/ RRID:SCR_021142 Blood vessel processing code This paper https://doi.org/10.5281/zenodo.14876892 GraphPad Prism 10.4.1 http://www.graphpad.com/ RRID:SCR_002798

    SYBR Green Assay:

    Article Title: Specific targeting of brain endothelial cells using enhancer AAV vectors.
    Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Ethidium Bromide (for maxiprep plasmids) Sigma CAS #: 1239-45-8 Isopropanol (for maxiprep plasmids) Fisher Scientific CAS #: 67-63-0 Phenol (for maxiprep plasmids) Fisher Scientific CAS #: 108-95-2 Chloroform (for maxiprep plasmids) Fisher Scientific CAS #: 67-66-3 Critical commercial assays Pierce Silver Stain Kit (AAV purification) Thermo Fisher Cat#: 24612 Deposited data CATlas http://catlas.org/mousebrain/#! .. RRID:SCR_018690 Mendeley Data: quantification and metadata This paper https://doi.org/10.17632/5kvfknrgng.1 UCSC Genome Browser http://genome.ucsc.edu/ RRID:SCR_005780 Experimental models: Cell lines Cells: AAVpro 293T (for virus packaging) Takara Bio RRID:CVCL_B0XW Experimental models: Organisms/strains Mouse: Tie2-Cre (B6.Cg-Tg(Tek-cre)1Ywa/J) The Jackson Laboratory RRID:IMSR_JAX:008863 Mouse: Ai9 (B6.Cg-Gt(ROSA)26Sortm9 (CAG-tdTomato)Hze/J) The Jackson Laboratory RRID:IMSR_JAX:007909 Mouse: 5xFAD (B6.Cg-Tg (APPSwFlLon,PSEN1*M146L*L286V)6799Vas/Mmjax) The Jackson Laboratory RRID:MMRRC_034848-JAX Mouse: B6J (C57BL/6J) The Jackson Laboratory RRID:IMSR_JAX:000664 Oligonucleotides Fwd ITR primer, 50-GGAACCCCTAGTGATGGAGTT Aurnhammer et al.68 PMID: 22428977 Rev ITR primer, 50-CGGCCTCAGTGAGCGA Aurnhammer et al.68 PMID: 22428977 PowerUP SYBR green Master mix Thermo Fisher A46012 Recombinant DNA pUCmini-iCAP-PHP.eB plasmid (encodes engineered PHP.eB capsid protein) Addgene Cat#: 103005; RRID:Addgene_103005 pAAV2/8 (encodes AAV8 capsid protein) Addgene Cat#: 112864; RRID:Addgene_112864 pAAV-BR1 Körbelin et al.13 PMID: 27137490 Ad helper plasmid (mini-pHelper) This paper N/A pAAV-BEC enhancer-minBG-SYFP2 This paper Cat #: 236722; RRID:Addgene_236722 pAAV-minBG-SYFP2 This paper Cat #: 236723; RRID:Addgene_236723 pAAV-CAG-mTurquoise2 Addgene Cat#: 99122; RRID:Addgene_99122 pAAV-BEC enhancer-minBG-iCre This paper Cat #: 236724; RRID:Addgene_236724 pAAV-BEC enhancer-minBG-iCre(R279T)_4x6T This paper Cat #: 236727; RRID:Addgene_236727 P3-Alb-mScarlet Addgene Cat#: 183461; RRID:Addgene_183461 AiP14187 - pAAV-AiE0459m_3xC2-minBGiCre(R297T)-BGHpA (plasmid used to clone out attenuated iCre(R279T)) Addgene Cat#: 229920; RRID:Addgene_229920 AAV-GfaABC1D-ERCreER-4x6T (plasmid used to clone out 4x6T) Addgene Cat#: 196411; RRID:Addgene_196411 Software and algorithms ImageJ (v. 1.53c) https://imagej.net/ RRID:SCR_003070 Python Programming Language http://www.python.org/ RRID:SCR_008394 OpenCV http://opencv.org RRID:SCR_015526 MatPlotLib http://matplotlib.sourceforge.net RRID:SCR_008624 NumPy http://www.numpy.org RRID:SCR_008633 SimpleITK https://simpleitk.org/ RRID:SCR_024693 (Continued on next page) e2 Neuron 113, 1562–1578.e1–e6, May 21, 2025 .. REAGENT or RESOURCE SOURCE IDENTIFIER SciPy http://www.scipy.org/ RRID:SCR_008058 Insight Segmentation and Registration Toolkit http://www.itk.org RRID:SCR_001149 scikit-image https://scikit-image.org/ RRID:SCR_021142 Blood vessel processing code This paper https://doi.org/10.5281/zenodo.14876892 GraphPad Prism 10.4.1 http://www.graphpad.com/ RRID:SCR_002798

    Recombinant:

    Article Title: Specific targeting of brain endothelial cells using enhancer AAV vectors.
    Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Ethidium Bromide (for maxiprep plasmids) Sigma CAS #: 1239-45-8 Isopropanol (for maxiprep plasmids) Fisher Scientific CAS #: 67-63-0 Phenol (for maxiprep plasmids) Fisher Scientific CAS #: 108-95-2 Chloroform (for maxiprep plasmids) Fisher Scientific CAS #: 67-66-3 Critical commercial assays Pierce Silver Stain Kit (AAV purification) Thermo Fisher Cat#: 24612 Deposited data CATlas http://catlas.org/mousebrain/#! .. RRID:SCR_018690 Mendeley Data: quantification and metadata This paper https://doi.org/10.17632/5kvfknrgng.1 UCSC Genome Browser http://genome.ucsc.edu/ RRID:SCR_005780 Experimental models: Cell lines Cells: AAVpro 293T (for virus packaging) Takara Bio RRID:CVCL_B0XW Experimental models: Organisms/strains Mouse: Tie2-Cre (B6.Cg-Tg(Tek-cre)1Ywa/J) The Jackson Laboratory RRID:IMSR_JAX:008863 Mouse: Ai9 (B6.Cg-Gt(ROSA)26Sortm9 (CAG-tdTomato)Hze/J) The Jackson Laboratory RRID:IMSR_JAX:007909 Mouse: 5xFAD (B6.Cg-Tg (APPSwFlLon,PSEN1*M146L*L286V)6799Vas/Mmjax) The Jackson Laboratory RRID:MMRRC_034848-JAX Mouse: B6J (C57BL/6J) The Jackson Laboratory RRID:IMSR_JAX:000664 Oligonucleotides Fwd ITR primer, 50-GGAACCCCTAGTGATGGAGTT Aurnhammer et al.68 PMID: 22428977 Rev ITR primer, 50-CGGCCTCAGTGAGCGA Aurnhammer et al.68 PMID: 22428977 PowerUP SYBR green Master mix Thermo Fisher A46012 Recombinant DNA pUCmini-iCAP-PHP.eB plasmid (encodes engineered PHP.eB capsid protein) Addgene Cat#: 103005; RRID:Addgene_103005 pAAV2/8 (encodes AAV8 capsid protein) Addgene Cat#: 112864; RRID:Addgene_112864 pAAV-BR1 Körbelin et al.13 PMID: 27137490 Ad helper plasmid (mini-pHelper) This paper N/A pAAV-BEC enhancer-minBG-SYFP2 This paper Cat #: 236722; RRID:Addgene_236722 pAAV-minBG-SYFP2 This paper Cat #: 236723; RRID:Addgene_236723 pAAV-CAG-mTurquoise2 Addgene Cat#: 99122; RRID:Addgene_99122 pAAV-BEC enhancer-minBG-iCre This paper Cat #: 236724; RRID:Addgene_236724 pAAV-BEC enhancer-minBG-iCre(R279T)_4x6T This paper Cat #: 236727; RRID:Addgene_236727 P3-Alb-mScarlet Addgene Cat#: 183461; RRID:Addgene_183461 AiP14187 - pAAV-AiE0459m_3xC2-minBGiCre(R297T)-BGHpA (plasmid used to clone out attenuated iCre(R279T)) Addgene Cat#: 229920; RRID:Addgene_229920 AAV-GfaABC1D-ERCreER-4x6T (plasmid used to clone out 4x6T) Addgene Cat#: 196411; RRID:Addgene_196411 Software and algorithms ImageJ (v. 1.53c) https://imagej.net/ RRID:SCR_003070 Python Programming Language http://www.python.org/ RRID:SCR_008394 OpenCV http://opencv.org RRID:SCR_015526 MatPlotLib http://matplotlib.sourceforge.net RRID:SCR_008624 NumPy http://www.numpy.org RRID:SCR_008633 SimpleITK https://simpleitk.org/ RRID:SCR_024693 (Continued on next page) e2 Neuron 113, 1562–1578.e1–e6, May 21, 2025 .. REAGENT or RESOURCE SOURCE IDENTIFIER SciPy http://www.scipy.org/ RRID:SCR_008058 Insight Segmentation and Registration Toolkit http://www.itk.org RRID:SCR_001149 scikit-image https://scikit-image.org/ RRID:SCR_021142 Blood vessel processing code This paper https://doi.org/10.5281/zenodo.14876892 GraphPad Prism 10.4.1 http://www.graphpad.com/ RRID:SCR_002798

    Plasmid Preparation:

    Article Title: Specific targeting of brain endothelial cells using enhancer AAV vectors.
    Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Ethidium Bromide (for maxiprep plasmids) Sigma CAS #: 1239-45-8 Isopropanol (for maxiprep plasmids) Fisher Scientific CAS #: 67-63-0 Phenol (for maxiprep plasmids) Fisher Scientific CAS #: 108-95-2 Chloroform (for maxiprep plasmids) Fisher Scientific CAS #: 67-66-3 Critical commercial assays Pierce Silver Stain Kit (AAV purification) Thermo Fisher Cat#: 24612 Deposited data CATlas http://catlas.org/mousebrain/#! .. RRID:SCR_018690 Mendeley Data: quantification and metadata This paper https://doi.org/10.17632/5kvfknrgng.1 UCSC Genome Browser http://genome.ucsc.edu/ RRID:SCR_005780 Experimental models: Cell lines Cells: AAVpro 293T (for virus packaging) Takara Bio RRID:CVCL_B0XW Experimental models: Organisms/strains Mouse: Tie2-Cre (B6.Cg-Tg(Tek-cre)1Ywa/J) The Jackson Laboratory RRID:IMSR_JAX:008863 Mouse: Ai9 (B6.Cg-Gt(ROSA)26Sortm9 (CAG-tdTomato)Hze/J) The Jackson Laboratory RRID:IMSR_JAX:007909 Mouse: 5xFAD (B6.Cg-Tg (APPSwFlLon,PSEN1*M146L*L286V)6799Vas/Mmjax) The Jackson Laboratory RRID:MMRRC_034848-JAX Mouse: B6J (C57BL/6J) The Jackson Laboratory RRID:IMSR_JAX:000664 Oligonucleotides Fwd ITR primer, 50-GGAACCCCTAGTGATGGAGTT Aurnhammer et al.68 PMID: 22428977 Rev ITR primer, 50-CGGCCTCAGTGAGCGA Aurnhammer et al.68 PMID: 22428977 PowerUP SYBR green Master mix Thermo Fisher A46012 Recombinant DNA pUCmini-iCAP-PHP.eB plasmid (encodes engineered PHP.eB capsid protein) Addgene Cat#: 103005; RRID:Addgene_103005 pAAV2/8 (encodes AAV8 capsid protein) Addgene Cat#: 112864; RRID:Addgene_112864 pAAV-BR1 Körbelin et al.13 PMID: 27137490 Ad helper plasmid (mini-pHelper) This paper N/A pAAV-BEC enhancer-minBG-SYFP2 This paper Cat #: 236722; RRID:Addgene_236722 pAAV-minBG-SYFP2 This paper Cat #: 236723; RRID:Addgene_236723 pAAV-CAG-mTurquoise2 Addgene Cat#: 99122; RRID:Addgene_99122 pAAV-BEC enhancer-minBG-iCre This paper Cat #: 236724; RRID:Addgene_236724 pAAV-BEC enhancer-minBG-iCre(R279T)_4x6T This paper Cat #: 236727; RRID:Addgene_236727 P3-Alb-mScarlet Addgene Cat#: 183461; RRID:Addgene_183461 AiP14187 - pAAV-AiE0459m_3xC2-minBGiCre(R297T)-BGHpA (plasmid used to clone out attenuated iCre(R279T)) Addgene Cat#: 229920; RRID:Addgene_229920 AAV-GfaABC1D-ERCreER-4x6T (plasmid used to clone out 4x6T) Addgene Cat#: 196411; RRID:Addgene_196411 Software and algorithms ImageJ (v. 1.53c) https://imagej.net/ RRID:SCR_003070 Python Programming Language http://www.python.org/ RRID:SCR_008394 OpenCV http://opencv.org RRID:SCR_015526 MatPlotLib http://matplotlib.sourceforge.net RRID:SCR_008624 NumPy http://www.numpy.org RRID:SCR_008633 SimpleITK https://simpleitk.org/ RRID:SCR_024693 (Continued on next page) e2 Neuron 113, 1562–1578.e1–e6, May 21, 2025 .. REAGENT or RESOURCE SOURCE IDENTIFIER SciPy http://www.scipy.org/ RRID:SCR_008058 Insight Segmentation and Registration Toolkit http://www.itk.org RRID:SCR_001149 scikit-image https://scikit-image.org/ RRID:SCR_021142 Blood vessel processing code This paper https://doi.org/10.5281/zenodo.14876892 GraphPad Prism 10.4.1 http://www.graphpad.com/ RRID:SCR_002798

    Bioprocessing:

    Article Title: Specific targeting of brain endothelial cells using enhancer AAV vectors.
    Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Ethidium Bromide (for maxiprep plasmids) Sigma CAS #: 1239-45-8 Isopropanol (for maxiprep plasmids) Fisher Scientific CAS #: 67-63-0 Phenol (for maxiprep plasmids) Fisher Scientific CAS #: 108-95-2 Chloroform (for maxiprep plasmids) Fisher Scientific CAS #: 67-66-3 Critical commercial assays Pierce Silver Stain Kit (AAV purification) Thermo Fisher Cat#: 24612 Deposited data CATlas http://catlas.org/mousebrain/#! .. RRID:SCR_018690 Mendeley Data: quantification and metadata This paper https://doi.org/10.17632/5kvfknrgng.1 UCSC Genome Browser http://genome.ucsc.edu/ RRID:SCR_005780 Experimental models: Cell lines Cells: AAVpro 293T (for virus packaging) Takara Bio RRID:CVCL_B0XW Experimental models: Organisms/strains Mouse: Tie2-Cre (B6.Cg-Tg(Tek-cre)1Ywa/J) The Jackson Laboratory RRID:IMSR_JAX:008863 Mouse: Ai9 (B6.Cg-Gt(ROSA)26Sortm9 (CAG-tdTomato)Hze/J) The Jackson Laboratory RRID:IMSR_JAX:007909 Mouse: 5xFAD (B6.Cg-Tg (APPSwFlLon,PSEN1*M146L*L286V)6799Vas/Mmjax) The Jackson Laboratory RRID:MMRRC_034848-JAX Mouse: B6J (C57BL/6J) The Jackson Laboratory RRID:IMSR_JAX:000664 Oligonucleotides Fwd ITR primer, 50-GGAACCCCTAGTGATGGAGTT Aurnhammer et al.68 PMID: 22428977 Rev ITR primer, 50-CGGCCTCAGTGAGCGA Aurnhammer et al.68 PMID: 22428977 PowerUP SYBR green Master mix Thermo Fisher A46012 Recombinant DNA pUCmini-iCAP-PHP.eB plasmid (encodes engineered PHP.eB capsid protein) Addgene Cat#: 103005; RRID:Addgene_103005 pAAV2/8 (encodes AAV8 capsid protein) Addgene Cat#: 112864; RRID:Addgene_112864 pAAV-BR1 Körbelin et al.13 PMID: 27137490 Ad helper plasmid (mini-pHelper) This paper N/A pAAV-BEC enhancer-minBG-SYFP2 This paper Cat #: 236722; RRID:Addgene_236722 pAAV-minBG-SYFP2 This paper Cat #: 236723; RRID:Addgene_236723 pAAV-CAG-mTurquoise2 Addgene Cat#: 99122; RRID:Addgene_99122 pAAV-BEC enhancer-minBG-iCre This paper Cat #: 236724; RRID:Addgene_236724 pAAV-BEC enhancer-minBG-iCre(R279T)_4x6T This paper Cat #: 236727; RRID:Addgene_236727 P3-Alb-mScarlet Addgene Cat#: 183461; RRID:Addgene_183461 AiP14187 - pAAV-AiE0459m_3xC2-minBGiCre(R297T)-BGHpA (plasmid used to clone out attenuated iCre(R279T)) Addgene Cat#: 229920; RRID:Addgene_229920 AAV-GfaABC1D-ERCreER-4x6T (plasmid used to clone out 4x6T) Addgene Cat#: 196411; RRID:Addgene_196411 Software and algorithms ImageJ (v. 1.53c) https://imagej.net/ RRID:SCR_003070 Python Programming Language http://www.python.org/ RRID:SCR_008394 OpenCV http://opencv.org RRID:SCR_015526 MatPlotLib http://matplotlib.sourceforge.net RRID:SCR_008624 NumPy http://www.numpy.org RRID:SCR_008633 SimpleITK https://simpleitk.org/ RRID:SCR_024693 (Continued on next page) e2 Neuron 113, 1562–1578.e1–e6, May 21, 2025 .. REAGENT or RESOURCE SOURCE IDENTIFIER SciPy http://www.scipy.org/ RRID:SCR_008058 Insight Segmentation and Registration Toolkit http://www.itk.org RRID:SCR_001149 scikit-image https://scikit-image.org/ RRID:SCR_021142 Blood vessel processing code This paper https://doi.org/10.5281/zenodo.14876892 GraphPad Prism 10.4.1 http://www.graphpad.com/ RRID:SCR_002798



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    encodes aav8 capsid protein  (Addgene inc)
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    Addgene inc encodes aav8 capsid protein
    Figure 5. BEC-enhancer AAV labels brain endothelial cells in brain blood vessels in vivo (A) Schematic experimental design to visualize endothelial cells in vivo. Mice received retro-orbital injection of BEC-enhancer AAV and <t>AAV8-P3-Alb-mScarlet</t> to label albumin in the blood and visualize brain blood vessels in vivo. The graphics were created with BioRender.com. (B) Representative in vivo two-photon imaging of anesthetized mice 4 weeks after AAV injection. Top left panel with 1-s averaged pixel density from blood albumin signal detected at single plane from somatosensory cortex in vivo. Top right panel with 1-s averaged pixel density from brain endothelial cells labeled by AAV- PHP.eB-BEC enhancer-minBG-SYFP2 from the same plane as the top left panel. Bottom, the merged image from both panels (see Video S2). (C) Representative in vivo two-photon imaging of smaller blood vessels relative to those shown in (B), detected for both BEC-enhancer AAV and AAV8-P3-Alb- mScarlet signals. (D) Representative in vivo two-photon imaging of anesthetized mice 10 weeks after AAV injection. Top panel with 5-s averaged pixel density from blood albumin signal depicted in magenta at a single plane from hippocampal CA1 in vivo. Middle panel with 5-s averaged pixel density from brain endothelial cells labeled by AAV-PHP.eB-BEC enhancer-minBG-SYFP2 (BEC-enhancer AAV, green) from the same plane as the top left panel. Bottom, the merged image of the above panels. The albumin signal as well as the signal from transduced cells with BEC-enhancer AAV is stable for at least a couple of months. Some speckles are due to debris artifacts.
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    Figure 5. BEC-enhancer AAV labels brain endothelial cells in brain blood vessels in vivo (A) Schematic experimental design to visualize endothelial cells in vivo. Mice received retro-orbital injection of BEC-enhancer AAV and AAV8-P3-Alb-mScarlet to label albumin in the blood and visualize brain blood vessels in vivo. The graphics were created with BioRender.com. (B) Representative in vivo two-photon imaging of anesthetized mice 4 weeks after AAV injection. Top left panel with 1-s averaged pixel density from blood albumin signal detected at single plane from somatosensory cortex in vivo. Top right panel with 1-s averaged pixel density from brain endothelial cells labeled by AAV- PHP.eB-BEC enhancer-minBG-SYFP2 from the same plane as the top left panel. Bottom, the merged image from both panels (see Video S2). (C) Representative in vivo two-photon imaging of smaller blood vessels relative to those shown in (B), detected for both BEC-enhancer AAV and AAV8-P3-Alb- mScarlet signals. (D) Representative in vivo two-photon imaging of anesthetized mice 10 weeks after AAV injection. Top panel with 5-s averaged pixel density from blood albumin signal depicted in magenta at a single plane from hippocampal CA1 in vivo. Middle panel with 5-s averaged pixel density from brain endothelial cells labeled by AAV-PHP.eB-BEC enhancer-minBG-SYFP2 (BEC-enhancer AAV, green) from the same plane as the top left panel. Bottom, the merged image of the above panels. The albumin signal as well as the signal from transduced cells with BEC-enhancer AAV is stable for at least a couple of months. Some speckles are due to debris artifacts.

    Journal: Neuron

    Article Title: Specific targeting of brain endothelial cells using enhancer AAV vectors.

    doi: 10.1016/j.neuron.2025.03.031

    Figure Lengend Snippet: Figure 5. BEC-enhancer AAV labels brain endothelial cells in brain blood vessels in vivo (A) Schematic experimental design to visualize endothelial cells in vivo. Mice received retro-orbital injection of BEC-enhancer AAV and AAV8-P3-Alb-mScarlet to label albumin in the blood and visualize brain blood vessels in vivo. The graphics were created with BioRender.com. (B) Representative in vivo two-photon imaging of anesthetized mice 4 weeks after AAV injection. Top left panel with 1-s averaged pixel density from blood albumin signal detected at single plane from somatosensory cortex in vivo. Top right panel with 1-s averaged pixel density from brain endothelial cells labeled by AAV- PHP.eB-BEC enhancer-minBG-SYFP2 from the same plane as the top left panel. Bottom, the merged image from both panels (see Video S2). (C) Representative in vivo two-photon imaging of smaller blood vessels relative to those shown in (B), detected for both BEC-enhancer AAV and AAV8-P3-Alb- mScarlet signals. (D) Representative in vivo two-photon imaging of anesthetized mice 10 weeks after AAV injection. Top panel with 5-s averaged pixel density from blood albumin signal depicted in magenta at a single plane from hippocampal CA1 in vivo. Middle panel with 5-s averaged pixel density from brain endothelial cells labeled by AAV-PHP.eB-BEC enhancer-minBG-SYFP2 (BEC-enhancer AAV, green) from the same plane as the top left panel. Bottom, the merged image of the above panels. The albumin signal as well as the signal from transduced cells with BEC-enhancer AAV is stable for at least a couple of months. Some speckles are due to debris artifacts.

    Article Snippet: RRID:SCR_018690 Mendeley Data: quantification and metadata This paper https://doi.org/10.17632/5kvfknrgng.1 UCSC Genome Browser http://genome.ucsc.edu/ RRID:SCR_005780 Experimental models: Cell lines Cells: AAVpro 293T (for virus packaging) Takara Bio RRID:CVCL_B0XW Experimental models: Organisms/strains Mouse: Tie2-Cre (B6.Cg-Tg(Tek-cre)1Ywa/J) The Jackson Laboratory RRID:IMSR_JAX:008863 Mouse: Ai9 (B6.Cg-Gt(ROSA)26Sortm9 (CAG-tdTomato)Hze/J) The Jackson Laboratory RRID:IMSR_JAX:007909 Mouse: 5xFAD (B6.Cg-Tg (APPSwFlLon,PSEN1*M146L*L286V)6799Vas/Mmjax) The Jackson Laboratory RRID:MMRRC_034848-JAX Mouse: B6J (C57BL/6J) The Jackson Laboratory RRID:IMSR_JAX:000664 Oligonucleotides Fwd ITR primer, 50-GGAACCCCTAGTGATGGAGTT Aurnhammer et al.68 PMID: 22428977 Rev ITR primer, 50-CGGCCTCAGTGAGCGA Aurnhammer et al.68 PMID: 22428977 PowerUP SYBR green Master mix Thermo Fisher A46012 Recombinant DNA pUCmini-iCAP-PHP.eB plasmid (encodes engineered PHP.eB capsid protein) Addgene Cat#: 103005; RRID:Addgene_103005 pAAV2/8 (encodes AAV8 capsid protein) Addgene Cat#: 112864; RRID:Addgene_112864 pAAV-BR1 Körbelin et al.13 PMID: 27137490 Ad helper plasmid (mini-pHelper) This paper N/A pAAV-BEC enhancer-minBG-SYFP2 This paper Cat #: 236722; RRID:Addgene_236722 pAAV-minBG-SYFP2 This paper Cat #: 236723; RRID:Addgene_236723 pAAV-CAG-mTurquoise2 Addgene Cat#: 99122; RRID:Addgene_99122 pAAV-BEC enhancer-minBG-iCre This paper Cat #: 236724; RRID:Addgene_236724 pAAV-BEC enhancer-minBG-iCre(R279T)_4x6T This paper Cat #: 236727; RRID:Addgene_236727 P3-Alb-mScarlet Addgene Cat#: 183461; RRID:Addgene_183461 AiP14187 - pAAV-AiE0459m_3xC2-minBGiCre(R297T)-BGHpA (plasmid used to clone out attenuated iCre(R279T)) Addgene Cat#: 229920; RRID:Addgene_229920 AAV-GfaABC1D-ERCreER-4x6T (plasmid used to clone out 4x6T) Addgene Cat#: 196411; RRID:Addgene_196411 Software and algorithms ImageJ (v. 1.53c) https://imagej.net/ RRID:SCR_003070 Python Programming Language http://www.python.org/ RRID:SCR_008394 OpenCV http://opencv.org RRID:SCR_015526 MatPlotLib http://matplotlib.sourceforge.net RRID:SCR_008624 NumPy http://www.numpy.org RRID:SCR_008633 SimpleITK https://simpleitk.org/ RRID:SCR_024693 (Continued on next page) e2 Neuron 113, 1562–1578.e1–e6, May 21, 2025

    Techniques: In Vivo, Injection, Imaging, Labeling