Cells were fixed with 4% paraformaldehyde for 20 mins, permeabilized and blocked with PBS with 0.3% Triton-X100 and 1% bovine serum albumin for 40 mins, then incubated with primary antibodies diluted with blocking solution at 4 °C overnight. Alexa Fluo series of second antibodies (Thermo Scientific) were applied accordingly for one hour at room temperature. Cells were finally mounted in 4′,6-diamidino-2-phenylindole (DAPI) and examined using fluorescence microscope (Leica DMi8). For BrdU-labeled cells, cells were fixed with cold methanol for 15 mins on ice, rehydrated with PBS for 10 mins, incubated with 2 M hydrochloric acid for 45 mins and neutralized with sodium borate for 15 mins before first antibody incubation. The first antibodies include rabbit anti-active caspase 3 (a-cas3), rabbit anti-BNP, rabbit anti-ANP, mouse anti-sarcomeric α-actinin (SAA; all from Abcam), rabbit anti- discoidin domain-containing receptor 2 (DDR2; Immunoway), mouse anti-smooth muscle actin (SMA; Cell Signaling Technology), Lectin from tomato, FITC conjugate (Sigma), rabbit anti-BrdU (Abcam), and rabbit anti-GATA binding protein 4 (GATA4; Proteintech).
Alexa fluo series of second antibodies
Manufactured by Thermo Fisher Scientific
The Alexa Fluor series of secondary antibodies are fluorescent-labeled antibodies used in various immunodetection techniques, such as immunofluorescence, flow cytometry, and Western blotting. They are designed to recognize and bind to primary antibodies, allowing for the detection and visualization of target proteins or cellular components. The Alexa Fluor dyes offer a range of emission wavelengths, enabling flexible experimental design and multicolor applications.
Automatically generated - may contain errors
Lab products found in correlation
Rabbit anti brdu, by Abcam (1 mentions)
Rabbit anti anp, by Abcam (1 mentions)
Fitc conjugate, by Merck Group (1 mentions)
Mouse anti gfap, by Merck Group (1 mentions)
Rabbit anti ki67, by Abcam (1 mentions)
Rabbit anti iba1, by Fujifilm (1 mentions)
Mouse anti th, by Merck Group (1 mentions)
Rabbit anti mecp2, by Cell Signaling Technology (1 mentions)
Rabbit anti nurr1, by Santa Cruz Biotechnology (1 mentions)
3 protocols using alexa fluo series of second antibodies
1
Multicolor Immunofluorescence Staining Protocol
2
LDH Cytotoxicity Assay and Immunostaining
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The LDH assay was performed according to manufactor's instructions (Beyotime) to measure the cytotoxicity in glial cells. Cytotoxicity was measured by subtracting LDH content in HDACitreated cells from total LDH in untreated controls. Immunofluorescence staining and imaging Cells were fixed with 4% paraformaldehyde for 20 minutes, permeabilized by 0.3% Triton-X100 and blocked by 1% bovine serum albumin for 40 minutes, then incubated with primary antibodies at 4 overnight. Alexa Fluo series of second antibodies (Thermo Scientific) were applied accordingly for one hour at room temperature. Cells were finally mounted in 4',6-diamidino-2phenylindole (DAPI) and examined using fluorescence microscope (Leica DMi8). The first antibodies include rabbit anti-Iba1 (Wako), mouse anti-GFAP (Sigma) and rabbit anti-Ki67 (Abcam).
3
Immunofluorescence Staining of Midbrain Cells
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Cells were fixed with 4% paraformaldehyde for 20 minutes, permeabilized and blocked with phosphate buffered saline with 0.3% Triton-X100 and 1% bovine serum albumin for 40 minutes, then incubated with first antibodies diluted with blocking solution at 4℃ overnight. Alexa Fluo series of second antibodies (Thermo Scientific)
were applied accordingly for two hours at room temperature. Cells were finally mounted in 4',6-diamidino-2-phenylindole (DAPI) and examined using fluorescence microscope (Leica DMi8). Embryonic and postnatal ventral midbrain tissues were fixed with 4% paraformaldehyde and dehydrated with 30% sucrose overnight, and cryosectioned at 14 μm thickness. The first antibodies used include rabbit anti-MeCP2
(Cell Signaling Technology), mouse anti-NURR1 (R&D Systems), mouse anti-TH (Sigma), rabbit anti-NURR1 (Santa Cruz Biotechnology).
were applied accordingly for two hours at room temperature. Cells were finally mounted in 4',6-diamidino-2-phenylindole (DAPI) and examined using fluorescence microscope (Leica DMi8). Embryonic and postnatal ventral midbrain tissues were fixed with 4% paraformaldehyde and dehydrated with 30% sucrose overnight, and cryosectioned at 14 μm thickness. The first antibodies used include rabbit anti-MeCP2
(Cell Signaling Technology), mouse anti-NURR1 (R&D Systems), mouse anti-TH (Sigma), rabbit anti-NURR1 (Santa Cruz Biotechnology).
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