To detect general anesthesia-activated neurons, animals were anesthetized with isoflurane, or ketamine/xylazine, or dexmedetomidine, for two hours then transcardially perfused with 10% sucrose in cold phosphate buffer saline (PBS, pH 7.4) followed by 4% cold paraformaldehyde (PFA) fixation solution. To detect restraint stress-activated neurons, mice were placed in the restrainer for 90 min (TV-150, Braintree scientific Inc). All brains were post-fixed in PFA overnight at 4°C, cryoprotected in 30% sucrose PBS solution for 2–3 days at 4°C, frozen in O.C.T compound (Tissue-Tek, Sakura), and then stored at −80°C until sectioning. Floating brain sections (80 µm) were stained using standard immunofluorescent protocol. The primary antibody used were: goat anti-Fos (Santa Cruz Biotechnology, sc520g, 1:300)50 , rabbit anti-Fos (Cell Signaling, #2250, 1:1500)51 , anti-Neurotensin (ImmunoStar, 20072)52 . The secondary antibody used are: Alexa Fluor 488 donkey anti-goat (Jackson ImmunoResearch, 705-545-147, 1:500)53 , Alexa Fluor 647 anti-rabbit (Jackson ImmunoResearch, 711-605-152, 1:500)54 , and Cy3 donkey anti-goat (Jackson ImmunoResearch, 705-165-147, 1:500)55 (see Life Sciences Reporting Summary).
Rabbit anti fos
Manufactured by Cell Signaling Technology
Rabbit anti-Fos is a primary antibody that recognizes the Fos protein, a transcription factor involved in cellular signaling pathways. It is a reliable tool for the detection and characterization of Fos expression in a variety of experimental systems.
Automatically generated - may contain errors
Lab products found in correlation
Alexa fluor 488 donkey anti goat, by Jackson ImmunoResearch (2 mentions)
Goat anti fos, by Santa Cruz Biotechnology (2 mentions)
Tv 150, by Braintree Scientific (2 mentions)
Anti neurotensin, by Immunostar (2 mentions)
Donkey anti goat cy3, by Jackson ImmunoResearch (2 mentions)
Alexa fluor 647 anti rabbit, by Jackson ImmunoResearch (2 mentions)
Rabbit anti pcreb, by Cell Signaling Technology (1 mentions)
Cm1850, by Leica (1 mentions)
Normal donkey serum (nds), by Jackson ImmunoResearch (1 mentions)
Alexa fluor 488 donkey anti rabbit, by Jackson ImmunoResearch (1 mentions)
Dapi fluoromount g, by Southern Biotech (1 mentions)
4 protocols using rabbit anti fos
1
Visualizing Stress-Induced Neural Activity
2
Immunofluorescence Analysis of pCREB and Fos in Basolateral Amygdala
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Mice were deeply anesthetized and were transcardially perfused with normal saline followed by 4 % paraformaldehyde in 0.1 M PB (pH 7.4). Brains were removed, postfixed in the same fixative overnight, and immersed in 20 % and 30 % sucrose (0.1 M PBS) for 24 ~ 48 h at 4 °C for cryoprotection. Coronal sections were cut at 30 μm with a cryostat (CM1850, Leica Microsystems). For immunofluorescence, free-floating sections were blocked by 5 % donkey serum in 0.01 M PBS with 0.3 % Triton-X100 for 1 h at RT. Sections were then incubated overnight at 4 °C with primary antibodies [rabbit anti- Fos (1:200, Cell Signaling) and rabbit anti-pCREB (1:1000)]. The sections were then incubated for 2 h at RT with Alexa Fluor 488-conjugated secondary antibody (donkey anti-rabbit, 1:200, Life Technologies). The sections were coverslipped, and then observed with a confocal laser scanning microscope (model FV1000, Olympus). For the quantification of immunoreactive signals, four non-adjacent sections from each mouse through the BLA were randomly selected. The numbers of pCREB- and Fos-labeled cells were counted in the BLA that was captured inside the optic field.
3
Visualizing Stress-Induced Neural Activity
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To detect general anesthesia-activated neurons, animals were anesthetized with isoflurane, or ketamine/xylazine, or dexmedetomidine, for two hours then transcardially perfused with 10% sucrose in cold phosphate buffer saline (PBS, pH 7.4) followed by 4% cold paraformaldehyde (PFA) fixation solution. To detect restraint stress-activated neurons, mice were placed in the restrainer for 90 min (TV-150, Braintree scientific Inc). All brains were post-fixed in PFA overnight at 4°C, cryoprotected in 30% sucrose PBS solution for 2–3 days at 4°C, frozen in O.C.T compound (Tissue-Tek, Sakura), and then stored at −80°C until sectioning. Floating brain sections (80 µm) were stained using standard immunofluorescent protocol. The primary antibody used were: goat anti-Fos (Santa Cruz Biotechnology, sc520g, 1:300)50 , rabbit anti-Fos (Cell Signaling, #2250, 1:1500)51 , anti-Neurotensin (ImmunoStar, 20072)52 . The secondary antibody used are: Alexa Fluor 488 donkey anti-goat (Jackson ImmunoResearch, 705-545-147, 1:500)53 , Alexa Fluor 647 anti-rabbit (Jackson ImmunoResearch, 711-605-152, 1:500)54 , and Cy3 donkey anti-goat (Jackson ImmunoResearch, 705-165-147, 1:500)55 (see Life Sciences Reporting Summary).
4
Immunohistochemical Labeling of c-Fos
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Brain sections were washed three times in PBS with 0.2% Triton X-100 (PBST) for 10 min at room temperature. Sections were then incubated in a blocking solution composed of PBST with 3% normal donkey serum (Jackson ImmunoResearch, #017-000-121) for 15 min at room temperature. For primary antibody exposure, sections were incubated in rabbit anti-Fos (1:1000; Cell Signaling Technology, #2250) in blocking solution overnight at 4 °C. After three 5 min washes in blocking solution, sections were incubated in AlexaFluor 488 donkey anti-rabbit (1:250; Jackson ImmunoResearch, #711-545-152) in block solution for 1 hr at room temperature. Finally, sections were washed three times in PBS. Following staining procedures, slides were coverslipped with DAPI Fluoromount-G (Southern Biotech, #0100–20) and stored in the dark at 4 °C until microscopy and imaging.
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