Immunohistochemical analysis was performed as previously described [25 ]. Mice were anesthetized at different time points after ICH induction, and frozen sections with the largest bleeding area were selected for immunofluorescence analysis. The following antibodies were used: mouse anti-GFAP (1:300; CST), mouse anti-Iba1 (1:250; GeneTex), mouse anti-NeuN (1:300; CST), rabbit anti-Homer1 (1:200; Abcam), rabbit anti-C3 (1:200; Abcam), rabbit anti-S100A10 (1:300; Proteintech), donkey anti-rabbit IgG (H + L) highly cross-adsorbed secondary antibody, Alexa Fluor Plus 488 (1:1000; Invitrogen), and donkey anti-mouse IgG (H + L) highly cross-adsorbed secondary antibody, Alexa Fluor Plus 555 (1:1000; Invitrogen).
Rabbit anti c3
Manufactured by Abcam
Sourced in United Kingdom
Rabbit anti-C3 is a polyclonal antibody raised against the C3 protein, a central component of the complement system. This antibody can be used to detect and analyze C3 in various research applications.
Automatically generated - may contain errors
Lab products found in correlation
Rabbit anti homer1, by Abcam (2 mentions)
Rabbit anti s100a10, by Proteintech (2 mentions)
Alexa fluor plus 488, by Thermo Fisher Scientific (1 mentions)
Alexa fluor plus 555, by Thermo Fisher Scientific (1 mentions)
Mouse anti iba1, by GeneTex (1 mentions)
Mouse anti gapdh, by Abcam (1 mentions)
Goat anti mouse rabbit secondary antibody, by Abcam (1 mentions)
Rabbit anti erk1 2, by Proteintech (1 mentions)
Rabbit anti gfap, by Abcam (1 mentions)
Alexa fluor 594, by Thermo Fisher Scientific (1 mentions)
Rat anti cd68, by Bio-Rad (1 mentions)
Goat anti gfap, by Abcam (1 mentions)
Rabbit anti iba1, by Fujifilm (1 mentions)
Alexa fluor 488, by Thermo Fisher Scientific (1 mentions)
Vibrance antifade mounting medium with dapi, by Vector Laboratories (1 mentions)
4 protocols using rabbit anti c3
1
Immunohistochemical Analysis of ICH in Mice
2
Western Blot Analysis of Astrocytic Proteins
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Western blotting was performed as previously described [25 ]. Primary astrocytes were harvested for protein extraction after the different treatments. Mice were anesthetized at different time points after ICH induction, and the brain tissue around the bleeding site was used for WB. The following antibodies were used: rabbit anti-Homer1 (1:1000; Abcam), rabbit anti-C3 (1:1000; Abcam), rabbit anti-S100A10 (1:1000; Proteintech), mouse anti-GAPDH (1:10,000; Abcam), rabbit anti-Ras (1:1000; CST), rabbit anti-Phospho-c-Raf (Ser338) (1:1000; CST), mouse anti-Raf-1 (1:1000; Proteintech), rabbit anti-Phospho-MEK1/2 (Ser217/221) (1:1000; CST), mouse anti-MEK1/2 (1:1000; CST), rabbit anti-Phospho-ERK1/2 (Thr202/Tyr204) (1:1000; Proteintech), rabbit anti-ERK1/2 (1:1000; Proteintech), and goat anti-mouse/rabbit secondary antibody (1:10,000; Abcam).
3
Immunofluorescence Staining of Astrocytes
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At the appropriate time, the cells were rinsed thrice with PBS and fixed in methanol at −20 °C for 15 min. Subsequently, they were blocked and permeabilized with 0.25 % Triton X-100 for 20 min. After further blocking in 3 % BSA for 1 h, they were incubated overnight with primary antibodies rabbit anti-GFAP (1:1000, Abcam), AQP4 (1:500, Proteintech) and rabbit anti-C3 (1:1000, Abcam) primary antibodies at 4 °C. The cells were subsequently, washed thrice with PBST, and incubated with an appropriate fluorescence-conjugated secondary antibody (1:1000, CST) at room temperature. Finally,the cells were stained with DAPI to visualize the nucleus. The images were acquired using fluorescence microscopy and analyzed with ImageJ.
4
Immunofluorescence of Retinal Cryosections
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Cryosections (12 μm) were incubated at room temperature in a blocking solution (5 % normal donkey serum, 0.3 % Triton X-100 in PBS) for 1 h. Subsequently, retinal sections were incubated with primary antibodies at 4 °C overnight. The following primary antibodies were used: rabbit anti-C3 (Abcam, Cambridge, UK), rabbit anti-IBA1 (Wako, Osaka, Japan), rat anti-CD68 (Bio-Rad, Hercules, CA, USA), goat anti-GFAP (Abcam), and mouse anti-FDP-lysine (ACR-modified protein; a gift from Dr. Uchida, University of Tokyo, Japan), mouse anti-GS (glutamine synthase; Abcam) antibody. After washing, sections were incubated with secondary antibodies conjugated to Alexa Fluor 488 or Alexa Fluor 594 (Invitrogen) for 1 h at room temperature. Sections were mounted using Vibrance Antifade Mounting Medium with DAPI (Vector Laboratories, Burlingame, CA, USA) and analyzed using BZ-X810.
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