Incubation:Article Title: GPR124 regulates hyaloid blood vessel regression and is associated with endothelial-mesenchymal transition
Article Snippet: .. Retinae were incubated with 5% normal goat serum (Thermo Fisher Scientific #31873) + 0.3% Triton X-100 in PBS (blocking buffer) for 24 h, followed by incubation with an Armenian hamster anti-CD31 antibody (1:1000, MilliporeSigma MAB1398Z) and a rabbit anti-collagen IV antibody (1:500, Bio-Rad #2150-1740) in blocking buffer for 48 h, both at 4 °C. .. Retinae were washed five times with PBS (20 min each) and incubated with a goat anti-Armenian hamster IgG-AF488 (1:1000, Jackson ImmunoResearch #127–545-160) and a goat anti-rabbit IgG-Cy3 secondary antibody (1:1000, Dianova/Biozol #112-165-003) at 4 °C for 24 h. After washing the tissue as before, retinae were transferred onto slides and radially incised four times at the periphery for flatmounting.
Article Title: Microbiome depletion rejuvenates the aging brain
Article Snippet: .. Sections were incubated overnight at 4 °C with primary antibodies diluted in blocking solution: goat anti-CD31 (1:100; R&D Systems, AF3628), rabbit anti-collagen IV (1:100; Bio-Rad, 2150-1470), mouse anti-ZO-1 (1:100; Thermo Fisher Scientific, 339100), mouse anti-Claudin-5 (1:100; Thermo Fisher Scientific, 352500), rat anti-CD68 (1:100; Bio-Rad, MCA1957), rabbit anti-Olig2 (Abcam, ab109186), chicken anti-MBP (Life Technologies, PA110008), rat anti-BrdU (1:100; Abcam, ab6326), mouse anti-NeuN (1:100; Millipore, MAB377), rabbit anti-Iba1 (1:100; FUJIFILM Wako, 01919741), and goat anti-Pdgfra (R&D Systems, AF1062). .. Nuclei were counterstained with DAPI (1:1000; Thermo Fisher Scientific, D1306).
Article Title: Vascular signals coordinate cerebellar circuitry development
Article Snippet: .. Primary antibodies: anti-Reelin (R&D, Af3820), anti-Pax6 (BioLegend, 901301 or Santa Cruz, Sc-81649), anti-Podocalyxin (R&D, AF1556), anti-phospho-Histone H3 (Merck, 06-570), anti-NeuN (Merck, MAB377 or ABN78), anti-p27 Kip1 (Cell Signaling, 83630), anti-Ki67 (Abcam, ab15580), anti-Wnt5a (LS Bio, LS-C124811), anti-Collagen 4 (Bio-Rad, 2150-1470), anti-Glut1 (Millipore, 07-1401), anti-Calbindin D-28k (Swant, 300, Merck, PC253L, Nitobe Medical, AB-2571569), anti-Vglut1(Merck, AB5905), anti-Vglut2 (Frontier Institute, AB_2571621), anti-Homer 1/2/3 (Synaptic Systems, 160 103), anti-Fzd2 (LSBio, LS-B3963) were diluted at 1:200 in 2% NDS, 0.1% Triton X-100 in PBS and incubated during 48 h at 4 °C. .. After consecutive washes in 0.1% Tween 20 in PBS, secondary antibodies: goat anti-Guinea pig A488 (Life Technologie, A11073), goat anti-rabbit A568 (Invitrogen, A32732), goat anti-mouse A647 (Invitrogen, A32728), donkey anti-mouse A488 (Invitrogen, A21202), A568 (Invitrogen, A10037), A647 (Invitrogen, A31571), donkey anti-rabbit A488 (Invitrogen, A21206), A555 (Invitrogen, A31572), A647 (Invitrogen, A31573), donkey anti-goat A488 (Invitrogen, A11055), A568 (Invitrogen, A11057), A647 (Invitrogen, A21447), donkey anti-rat A488 (Invitrogen, A21208), were diluted at 1:200 in the same solution and incubated overnight at 4 °C.
Blocking Assay:Article Title: GPR124 regulates hyaloid blood vessel regression and is associated with endothelial-mesenchymal transition
Article Snippet: .. Retinae were incubated with 5% normal goat serum (Thermo Fisher Scientific #31873) + 0.3% Triton X-100 in PBS (blocking buffer) for 24 h, followed by incubation with an Armenian hamster anti-CD31 antibody (1:1000, MilliporeSigma MAB1398Z) and a rabbit anti-collagen IV antibody (1:500, Bio-Rad #2150-1740) in blocking buffer for 48 h, both at 4 °C. .. Retinae were washed five times with PBS (20 min each) and incubated with a goat anti-Armenian hamster IgG-AF488 (1:1000, Jackson ImmunoResearch #127–545-160) and a goat anti-rabbit IgG-Cy3 secondary antibody (1:1000, Dianova/Biozol #112-165-003) at 4 °C for 24 h. After washing the tissue as before, retinae were transferred onto slides and radially incised four times at the periphery for flatmounting.
Article Title: Microbiome depletion rejuvenates the aging brain
Article Snippet: .. Sections were incubated overnight at 4 °C with primary antibodies diluted in blocking solution: goat anti-CD31 (1:100; R&D Systems, AF3628), rabbit anti-collagen IV (1:100; Bio-Rad, 2150-1470), mouse anti-ZO-1 (1:100; Thermo Fisher Scientific, 339100), mouse anti-Claudin-5 (1:100; Thermo Fisher Scientific, 352500), rat anti-CD68 (1:100; Bio-Rad, MCA1957), rabbit anti-Olig2 (Abcam, ab109186), chicken anti-MBP (Life Technologies, PA110008), rat anti-BrdU (1:100; Abcam, ab6326), mouse anti-NeuN (1:100; Millipore, MAB377), rabbit anti-Iba1 (1:100; FUJIFILM Wako, 01919741), and goat anti-Pdgfra (R&D Systems, AF1062). .. Nuclei were counterstained with DAPI (1:1000; Thermo Fisher Scientific, D1306).
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Isolation:Article Title: Exploiting porphyrin metabolism to inhibit angiogenesis
Article Snippet: .. Retinas were isolated at P16 for immunolabeling with antibodies anti collagen IV (2150-1470 Bio-Rad) and alpha smooth muscle actin (αSMA; C6198 Sigma-Aldrich) and images were acquired with a stereo fluorescent microscope (SZX16; Olympus) equipped with a digital camera (Hamamatsu Photonics). .. Artero-venous shunts were manually quantified, whereas central avascular (AV) and neovascular (NV) areas were measured with the open-source software ImageJ as previously described [ ].
Article Title: Exploiting porphyrin metabolism to inhibit angiogenesis.
Article Snippet: .. Retinas were isolated at P16 for immunolabeling with antibodies anti collagen IV (2150-1470 Bio-Rad) and alpha smooth muscle actin (αSMA; C6198 Sigma-Aldrich) and images were acquired with a stereo fluorescent microscope (SZX16; Olympus) equipped with a digital camera (Hamamatsu Photonics). .. Artero-venous shunts were manually quantified, whereas central avascular (AV) and neovascular (NV) areas were measured with the open-source software ImageJ as previously described [19].
Immunolabeling:Article Title: Exploiting porphyrin metabolism to inhibit angiogenesis
Article Snippet: .. Retinas were isolated at P16 for immunolabeling with antibodies anti collagen IV (2150-1470 Bio-Rad) and alpha smooth muscle actin (αSMA; C6198 Sigma-Aldrich) and images were acquired with a stereo fluorescent microscope (SZX16; Olympus) equipped with a digital camera (Hamamatsu Photonics). .. Artero-venous shunts were manually quantified, whereas central avascular (AV) and neovascular (NV) areas were measured with the open-source software ImageJ as previously described [ ].
Article Title: Exploiting porphyrin metabolism to inhibit angiogenesis.
Article Snippet: .. Retinas were isolated at P16 for immunolabeling with antibodies anti collagen IV (2150-1470 Bio-Rad) and alpha smooth muscle actin (αSMA; C6198 Sigma-Aldrich) and images were acquired with a stereo fluorescent microscope (SZX16; Olympus) equipped with a digital camera (Hamamatsu Photonics). .. Artero-venous shunts were manually quantified, whereas central avascular (AV) and neovascular (NV) areas were measured with the open-source software ImageJ as previously described [19].
Microscopy:Article Title: Exploiting porphyrin metabolism to inhibit angiogenesis
Article Snippet: .. Retinas were isolated at P16 for immunolabeling with antibodies anti collagen IV (2150-1470 Bio-Rad) and alpha smooth muscle actin (αSMA; C6198 Sigma-Aldrich) and images were acquired with a stereo fluorescent microscope (SZX16; Olympus) equipped with a digital camera (Hamamatsu Photonics). .. Artero-venous shunts were manually quantified, whereas central avascular (AV) and neovascular (NV) areas were measured with the open-source software ImageJ as previously described [ ].
Article Title: Exploiting porphyrin metabolism to inhibit angiogenesis.
Article Snippet: .. Retinas were isolated at P16 for immunolabeling with antibodies anti collagen IV (2150-1470 Bio-Rad) and alpha smooth muscle actin (αSMA; C6198 Sigma-Aldrich) and images were acquired with a stereo fluorescent microscope (SZX16; Olympus) equipped with a digital camera (Hamamatsu Photonics). .. Artero-venous shunts were manually quantified, whereas central avascular (AV) and neovascular (NV) areas were measured with the open-source software ImageJ as previously described [19].
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