The rats were killed by intraperitoneal injection (10% chloral hydrated, 1 ml/100 g) and 4% paraformaldehyde (PFA) was used to fix via cardiac perfusion. Coronal brain slices (15 μm thick) were prepared and used for staining. Slices were washed in PBS for 5 min, three times, then blotted in 5% goat serum for 1 h at room temperature. Slices were incubated with the primary antibodies overnight at 4°C. Primary antibodies used: mouse anti-Iba1 (1:200 Sigma American), mouse anti-GFAP (1:200 Sigma American), rabbit anti-iNOS, anti-Arg1, anti-IL-1β, anti-TNF-α, anti-IL-4, anti-IL-6, and anti-IL-10 (1:200; Bioss, Beijing, China). After having been washed in PBS three times, these slices were incubated with secondary antibodies at 37°C for 2 h. Secondary antibodies: Alexa Fluor 488-conjugated goat anti-rabbit IgG (1:200; Bioss, Beijing, China), Cy3-conjugated goat ant-mouse IgG (1:200; Bioss, Beijing, China). Then these slices were washed three times again and stained with DAPI (Biyuntian, China) for 5 min. Results were imaged using a 7266-fluorescence microscope (Leica, Japan).
Anti arg1
Manufactured by Bioss Antibodies
Sourced in China
Anti-Arg1 is a lab equipment product manufactured by Bioss Antibodies. It is a highly specific antibody that targets the Arginase-1 (Arg1) protein.
Automatically generated - may contain errors
Lab products found in correlation
Anti il 10, by Bioss Antibodies (3 mentions)
Rabbit anti inos, by Bioss Antibodies (3 mentions)
Anti il 4, by Bioss Antibodies (2 mentions)
Anti il 6, by Bioss Antibodies (2 mentions)
Anti il 1β, by Bioss Antibodies (2 mentions)
Anti tnf α, by Bioss Antibodies (2 mentions)
Alexa fluor 488 conjugated goat anti rabbit igg, by Bioss Antibodies (2 mentions)
7266 fluorescence microscope, by Leica (2 mentions)
4 6 diamidino 2 phenylindole (dapi), by Beyotime (1 mentions)
Apotome 2, by Zeiss (1 mentions)
Nis elements advanced research 4, by Nikon (1 mentions)
Anti mpo, by Abcam (1 mentions)
Anti ly6g, by Abcam (1 mentions)
Anti foxp3, by Cell Signaling Technology (1 mentions)
Anti f4 80, by BioLegend (1 mentions)
Anti cd206, by Abcam (1 mentions)
Pt link system, by Agilent Technologies (1 mentions)
Anti ki67, by Abcam (1 mentions)
Anti cd8, by Agilent Technologies (1 mentions)
Axio observer light microscope, by Zeiss (1 mentions)
5 protocols using anti arg1
1
Immunohistochemical Analysis of Neuroinflammation
2
Immune Profiling in Melanoma Metastasis
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Deparaffinization and antigen retrieval were performed in mouse melanoma lung metastasis using a PT-link system (Dako) and stained as previously described (13 (link)) The following antibodies were used for immune stainings: anti-iNOS, anti-CD206, anti-CD103, anti-Ki67, anti-granzyme B, anti-MPO, anti-CD86, anti-CD68, anti-MHC-II, anti-CD11b, anti-Ly6C, anti-Ly6G, and anti-PD-L1 all purchased from Abcam; anti-CD11c and anti-F4/80, purchased from BioLegend; anti-Foxp3 (Cell Signaling); anti-Arg1 (Bioss) and anti-CD8 (Dako) primary antibodies, anti-CD4 (BioLegend) and anti-CD25 (R&D Systems) followed by fluorescently labeled secondary antibodies. Images were acquired using an Axio Observer Light Microscope with the Apotome.2 (Zeiss). Metastatic melanoma lesions were gated by generating a region of interest (ROI), and threshold merge fluorescence was limited to ROI and calculated using the NIS-Elements Advanced Research 4.0 software (Nikon, Tokyo).
3
Immunofluorescence Assay for Microglial Markers
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Cells were fixed with 4% paraformaldehyde for 10 min and then washed in phosphate-buffered saline (PBS). After being blocked with 5% bovine serum albumin containing 0.5% Triton X-100 for 1 h at 37°C, the microglia were incubated with the following primary antibodies at 4°C overnight: mouse anti-Iba1 (1:200, Bioss, Beijing, China), rabbit anti-iNOS, anti-Arg1, anti-IL-1β, anti-TNF-α, anti-IL-4, anti-IL-6 and anti-IL-10 (1:200, Bioss). After being washed in PBS three times, the cells were incubated with the following secondary antibodies at 37°C for 1 h: Alexa Fluor 488-conjugated goat anti-rabbit IgG (1:200, Bioss) or Alexa Fluor Cy3-conjugated goat anti-mouse IgG (1:200, Bioss). After being washed in PBS three times, the cells were incubated with 4′,6-diamidino-2-phenylindole (DAPI; Biyuntian, China) for 5 min. The cells were then washed in PBS and observed under a 7266-fluorescence microscope (Leica, Japan).
4
Protein Extraction and Western Blot
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For protein extraction, 1 mL of NP-40 lysis buffer (Biyuntian) was transferred to each well of a six-well plate of microglia on ice for 30 min, and the cells were then centrifuged at 12,000 rpm for 10 min. The soluble protein solutions were then mixed with 5x sample buffer (Biyuntian) and boiled at 90oC for 10 min. Equal amounts of protein (60 μg) from each sample were separated via 10% sodium dodecyl sulfate polyacrylamide gel electrophoresis (Sigma) and transferred to polyvinylidene fluoride membranes (Millipore, Billerica, MA, USA). The membranes were blocked in 5% milk with Tris-buffered saline for 1 h, and then were incubated with the following primary antibodies at 4°C overnight: rabbit anti-iNOS, anti-Arg1 (1:500, Bioss) and rabbit anti-actin (1:1000, Santa Cruz Biotechnology). The membranes were washed three times in Tris-buffered saline with Tween and then incubated with the secondary antibody (peroxidase-conjugated IgG, 1:5000, Santa) at room temperature for 2 h. Binding antibodies were visualized using enhanced chemiluminescence on an imaging system (Amersham Imager 600).
5
Anti-Inflammatory and Anti-Fibrotic Mechanisms
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Paeoniflorin was purchased from Yuanye Bio-Technology (Shanghai, China), and lificiguat (YC-1) from SelleckChem (Houston, TX, USA). CCl 4 was provided by Tianjin Chemical (Tianjin, China), and CoCl 2 was obtained from Makclin Biochemical Co. Ltd. The antibodies targeting MMP2, P65, p-P65, p-I B / , and p-IKK were acquired from Cell Signaling Technologies (Danvers, MA, USA). The anti-MMP9 and anti-TGF-antibodies were from Abcam (Cambridge, MA, UK), and the anti-HIF-, anti-TNF-, and anti-IL-6 antibodies were from Affinity Biosciences (Jiangsu, China). The anti-F4/80, anti-iNOS, anti-ARG-1, anti-CD163, anti-IL-10, and anti-TIMP-1 antibodies were from Bioss Biotechnology (Beijing, China).
In addition, antibodies specific for -SMA, GAPDH, and -actin, as well as the HRP-linked goat, anti-rabbit, and goat antirat IgG antibodies were obtained from Servicebio (Wuhan, China). The rabbit immunohistochemistry kit was provided by ZSGB Biotechnology (Beijing, China), and Alexa Fluor 488/594 goat anti-rat/rabbit IgG H&L was purchased from Invitrogen (CA, USA).
In addition, antibodies specific for -SMA, GAPDH, and -actin, as well as the HRP-linked goat, anti-rabbit, and goat antirat IgG antibodies were obtained from Servicebio (Wuhan, China). The rabbit immunohistochemistry kit was provided by ZSGB Biotechnology (Beijing, China), and Alexa Fluor 488/594 goat anti-rat/rabbit IgG H&L was purchased from Invitrogen (CA, USA).
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