sheep anti trem2  (R&D Systems)

Journal: bioRxiv

Article Title: Apolipoprotein E ε4 exacerbates microglia-mediated complement-dependent synapse loss caused by neuronal Tpk deficiency

doi: 10.1101/2025.01.16.633468

Figure Lengend Snippet: a. Flow chart of injection of SB290157. The injection of SB290157 started from the 4 weeks after tamoxifen induction, the frequency of injections is 3 times per week. b. Scores of nest building. The results showed that the injection of SB290157 did not alleviate the abilities of nest building of the AE-cKO mice. N = 5 per group. c. Rotarod test. The results showed that the injection of SB290157 did not alleviate the run time on the rotarod of the AE-cKO mice. N = 5 per group. d. Y-maze test. The results showed that the injection of SB290157 did not alleviate the behavior abnormality of the AE-cKO mice. N = 5 per group. e-g. Representative images of Iba1 (red) immunostaining ( e ) and quantification ( f,g ) in the cortices of mice. The results showed that the injection of SB290157 significantly decreased the activation of microglia in the cortices of the AE-cKO mice, but did not affect the count of Iba. h, i. Representative images of Iba1 (red) and CD68 (green) immunostaining ( h ) and quantification ( i ) in the cortices of mice. The results showed that the injection of SB290157 significantly decreased the elevation of co-labeled cells stained by Iba1 and CD68 antibodies in the AE-cKO mice. j, k. Representative images of Iba1 (red) and TSPO (green) immunostaining ( j ) and quantification ( k ) in the cortices of mice. The results showed that the injection of SB290157 significantly downregulated the enhancement of co-labeled cells stained by Iba1 and TSPO antibodies in the AE-cKO mice. l, m. Representative images of Iba1 (red) and Trem2 (green) immunostaining ( l ) and quantification ( m ) in the cortices of mice. The results showed that the humanized APOE4 knock-in significantly decreased the elevation of co-labeled cells stained by Iba1 and Trem2 antibodies in the AE-cKO mice. N = 5 mice per group. n-p. Representative images of western blotting ( n ) and quantification ( o, p ) of synaptophysin and PSD95 in the cortices of mice. The results showed that the injection of SB290157 significantly alleviated the reduction of synaptophysin ( n, p ) and PSD95 ( n, o ) in the cortices of the AE-cKO mice. N = 6 mice per group. The data represent mean ± SEM. * p <0.05, ** p <0.01, *** p <0.001, **** p <0.0001, one-way ANOVA followed by Tukey’s post hoc test. Scale bars, 25 μm.

Article Snippet: The following primary antibodies were used: rat anti-Iba1 (1:500, ab283346, Abcam), goat anti-Iba1 (1:200, ab5076, Abcam), goat anti-GFAP (1:200, ab53554, Abcam), mouse anti-NeuN (1:200, MAB377, Millipore), rabbit anti-CD68 (1:200, ab283654, Abcam), sheep anti-trem2 (1:200, AF1729, R&D), rabbit anti-4G8 (1:200,800702, Bio Legend), rabbit anti-TSPO (1:2000, ab109497, Abcam), rat anti-C3 (1:200, ab11862, Abcam), and rabbit anti-C1q (1:200, ab182451,abcam).

Techniques: Injection, Immunostaining, Activation Assay, Labeling, Staining, Knock-In, Western Blot

Journal: bioRxiv

Article Title: Apolipoprotein E ε4 exacerbates microglia-mediated complement-dependent synapse loss caused by neuronal Tpk deficiency

doi: 10.1101/2025.01.16.633468

Figure Lengend Snippet: a, b. Representative images of Iba1 (green) immunostaining ( a ) and quantification ( b ) in the cortices of mice. The results showed that the humanized APOE4 knock-in significantly worsened the activation of microglia in the cortices induced by neuronal Tpk ablation, but did not affect those in the mice without Tpk knockout. c, d. Representative images of Iba1 (red) and TSPO (green) immunostaining ( c ) and quantification ( d ) in the cortices of mice. The results showed that the humanized APOE4 knock-in significantly upregulated the enhancement of co-labeled cells by Iba1 and TSPO antibodies in the cKO mice, but did not affect those in mice without Tpk knockout. e, f. Representative images of Iba1 (red) and CD68 (green) immunostaining ( e ) and quantification ( f ) in the cortices of mice. The results showed that the humanized APOE4 knock-in significantly exacerbated the elevation of co-labeled cells by Iba1 and CD68 antibodies in the cKO mice, but did affect those in mice without Tpk knockout. g, h. Representative images of Iba1 (green) and Trem2 (red) immunostaining ( g ) and quantification ( h ) in the cortices of mice. The results showed that the humanized APOE4 knock-in significantly aggravated the enhancement of co-labeled cells by Iba1 and Trem2 antibodies in the cKO mice but did not affect those in mice without Tpk knockout. N = 5 mice per group. The data represent mean ± SEM. * p <0.05, ** p <0.01, *** p <0.001, **** p <0.0001, one-way ANOVA followed by Tukey’s post hoc test. Scale bars, 25 μm. i Representative images of immunostaining of Iba1 (red) and PSD95 (green) i n the cortices of mice. The results showed that microglia-mediated synaptic phagocytosis presented by PSD95 in the microglia. Scale bars, 10 μ

Article Snippet: The following primary antibodies were used: rat anti-Iba1 (1:500, ab283346, Abcam), goat anti-Iba1 (1:200, ab5076, Abcam), goat anti-GFAP (1:200, ab53554, Abcam), mouse anti-NeuN (1:200, MAB377, Millipore), rabbit anti-CD68 (1:200, ab283654, Abcam), sheep anti-trem2 (1:200, AF1729, R&D), rabbit anti-4G8 (1:200,800702, Bio Legend), rabbit anti-TSPO (1:2000, ab109497, Abcam), rat anti-C3 (1:200, ab11862, Abcam), and rabbit anti-C1q (1:200, ab182451,abcam).

Techniques: Immunostaining, Knock-In, Activation Assay, Knock-Out, Labeling