mouse anti aβ 6e10, by Fortrea - Product details

1

Cortical Brain Tissue Analysis of APP-KI Mice

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The brain cortical tissues were collected from APP-KI mice with or without SR9009 injection. The brain lysates were prepared as described previously [23 (link)]. The following primary antibodies were used: rabbit anti-Iba1 (1:1000; WAKO), mouse anti-NOS2 (1:1000; Abcam), mouse anti-IL-1β (1:1000, Santa Cruz Biotechnology), mouse anti-actin (1:5000; Abcam), mouse anti-phospho-IκBα (1:1000, Santa Cruz Biotechnology), rabbit anti-IκBα (1:1000, Santa Cruz Biotechnology), and mouse anti-Aβ (6E10, 1:1000, Covance). The following were used as secondary antibodies: horseradish peroxidase (HRP)-labeled anti-rabbit (1:2000; GE Healthcare) and anti-mouse (1:2000; R&D Systems). The HRP-labeled antibodies were detected using an enhanced chemiluminescence detection system (ECL Kit; GE Healthcare) with an image analyzer (LAS-1000; Fuji Photo Film).

Ni J., Wu Z., Meng J., Saito T., Saido T.C., Qing H, & Nakanishi H. (2019). An impaired intrinsic microglial clock system induces neuroinflammatory alterations in the early stage of amyloid precursor protein knock-in mouse brain. Journal of Neuroinflammation, 16, 173.

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Western Blotting of Amyloid-Beta in Brain

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For Western Blotting, total brain homogenates were sonicated 3x5 seconds (LabSonic, B. Braun Biotech), protein levels of the brain homogenates were quantified with a microplate bicinchoninic acid (BCA) assay (Pierce) and adjusted accordingly. Samples were then analysed on NuPage Bis-Tris gels (Invitrogen) using standard procedures. Proteins were transferred to nitrocellulose membranes, blocking was performed with 5% milk in PBS containing 0.05% Tween (PBST) for 1h and blots were incubated with mouse anti-Aβ (6E10; 1:1000, Covance) in PBST overnight at 4°C. Membranes were then probed with the secondary HRP-labelled antibodies (1:20,000, Jackson ImmunoLaboratories). Protein bands were detected using chemiluminescent peroxidase substrate (ECL prime, GE Healthcare). Densitometric values of the protein band intensities were analysed with the software package Aida v.4.27 and normalised to GAPDH intensities.

Wendeln A.C., Degenhardt K., Kaurani L., Gertig M., Ulas T., Jain G., Wagner J., Häsler L.M., Wild K., Skodras A., Blank T., Staszewski O., Datta M., Centeno T.P., Capece V., Islam M.R., Kerimoglu C., Staufenbiel M., Schultze J.L., Beyer M., Prinz M., Jucker M., Fischer A, & Neher J.J. (2018). Innate immune memory in the brain shapes neurological disease hallmarks. Nature, 556(7701), 332-338.

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Immunofluorescent Staining of Mouse Brain Samples

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Mouse brain samples for immunofluorescent staining were prepared as previously reported [20 (link), 21 (link)]. Sections for staining were incubated with the following primary antibodies: rabbit anti Iba1 (1:2000, Wako) and mouse anti-Aβ (6E10, 1:1000, Covance) at 4 °C overnight. After being washing with PBS, the sections were incubated with donkey anti-rabbit Alexa 488 (1:500; Jackson ImmunoResearch) and donkey anti-mouse Cy3 (1:500; Jackson ImmunoResearch) at 4 °C for 2 h. After further washing, the sections were mounted in Vectashield anti-fading medium (Vector Laboratories). The fluorescence images were observed using a confocal laser scanning microscope (CLSM; C2si, Nikon).

Ni J., Wu Z., Meng J., Saito T., Saido T.C., Qing H, & Nakanishi H. (2019). An impaired intrinsic microglial clock system induces neuroinflammatory alterations in the early stage of amyloid precursor protein knock-in mouse brain. Journal of Neuroinflammation, 16, 173.

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Western Blotting of Amyloid-Beta in Brain

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For Western Blotting, total brain homogenates were sonicated 3x5 seconds (LabSonic, B. Braun Biotech), protein levels of the brain homogenates were quantified with a microplate bicinchoninic acid (BCA) assay (Pierce) and adjusted accordingly. Samples were then analysed on NuPage Bis-Tris gels (Invitrogen) using standard procedures. Proteins were transferred to nitrocellulose membranes, blocking was performed with 5% milk in PBS containing 0.05% Tween (PBST) for 1h and blots were incubated with mouse anti-Aβ (6E10; 1:1000, Covance) in PBST overnight at 4°C. Membranes were then probed with the secondary HRP-labelled antibodies (1:20,000, Jackson ImmunoLaboratories). Protein bands were detected using chemiluminescent peroxidase substrate (ECL prime, GE Healthcare). Densitometric values of the protein band intensities were analysed with the software package Aida v.4.27 and normalised to GAPDH intensities.

Wendeln A.C., Degenhardt K., Kaurani L., Gertig M., Ulas T., Jain G., Wagner J., Häsler L.M., Wild K., Skodras A., Blank T., Staszewski O., Datta M., Centeno T.P., Capece V., Islam M.R., Kerimoglu C., Staufenbiel M., Schultze J.L., Beyer M., Prinz M., Jucker M., Fischer A, & Neher J.J. (2018). Innate immune memory in the brain shapes neurological disease hallmarks. Nature, 556(7701), 332-338.

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Mouse anti aβ 6e10

Lab products found in correlation

4 protocols using mouse anti aβ 6e10

Cortical Brain Tissue Analysis of APP-KI Mice

Western Blotting of Amyloid-Beta in Brain

Immunofluorescent Staining of Mouse Brain Samples

Western Blotting of Amyloid-Beta in Brain

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