Free-floating sections were washed 3 times for 10 minutes with TBS with 0.2% Triton X-100 (Roche, Indianapolis, IN) and blocked in 5% Normal Goat Serum (NGS; Jackson ImmunoResearch, West Grove, PA) with 0.2% Triton X-100 in TBS for 1 hour at room temperature. Primary antibodies (see table below) were diluted in 5% NGS in TBS with 0.2% Triton X-100. Sections were incubated overnight at 4°C with primary antibodies and washed three times for 10 minutes with TBS the following morning. Secondary Alexa-fluorophore conjugated antibodies (Invitrogen, Carlsbad, CA) were diluted (1:200) in 5% NGS in TBS with 0.2% Triton X-100, and sections were incubated with secondary antibodies for 2 hours at room temperature, protected from light. After incubation, sections were washed three times for 15 minutes in TBS and mounted with VECTASHIELD with DAPI (Vector Laboratories, Burlingame, CA). Images were acquired on an Olympus FV3000 confocal laser-scanning microscope.
Alexa fluorophore conjugated antibodies
Manufactured by Thermo Fisher Scientific
Alexa-fluorophore conjugated antibodies are a type of labeling reagent used in various biological and biochemical applications. These antibodies are conjugated with Alexa fluorophores, which are a class of bright, photostable, and pH-insensitive fluorescent dyes. The core function of these labeled antibodies is to serve as detection agents, allowing the visualization and quantification of target molecules in various experimental techniques, such as immunofluorescence, flow cytometry, and Western blotting.
Automatically generated - may contain errors
Lab products found in correlation
Fv3000 confocal laser scanning microscope, by Olympus (12 mentions)
D1306, by Thermo Fisher Scientific (8 mentions)
Normal goat serum (ngs), by Jackson ImmunoResearch (6 mentions)
Vectashield with dapi, by Vector Laboratories (4 mentions)
Hoechst 33258, by Thermo Fisher Scientific (4 mentions)
Anti gfap, by Merck Group (3 mentions)
Lsm 710 confocal microscope, by Zeiss (3 mentions)
Normal donkey serum (nds), by Jackson ImmunoResearch (2 mentions)
Anti tuj1, by Fortrea (2 mentions)
Anti nestin, by BD (2 mentions)
Gt anti sox1, by R&D Systems (1 mentions)
Hrp linked antibodies, by GE Healthcare (1 mentions)
Vt1200, by Leica (1 mentions)
Anti dcx, by Abcam (1 mentions)
Anti hu nuclei, by Merck Group (1 mentions)
Anti dcx, by Merck Group (1 mentions)
Anti sox2, by Abcam (1 mentions)
Anti ki67, by Vector Laboratories (1 mentions)
Anti synaptophysin, by Merck Group (1 mentions)
Anti gaba, by Abcam (1 mentions)
18 protocols using alexa fluorophore conjugated antibodies
1
Immunofluorescence Staining Protocol for Tissue Sections
2
Immunofluorescence Labeling Protocol
3
Immunofluorescence Labeling of Brain Tissue
4
Immunofluorescence Staining Protocol
5
Immunohistochemistry Protocol for Neuronal and Immune Cell Markers
6
Immunofluorescence Staining Protocol for Tissue Sections
7
Pluripotency and Lineage Marker Antibody Panel
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The following primary antibodies were used for Immunofluorescence stainings: m anti TRIM32 (Abnova), rb anti TRIM32 (Gramsch Laboratories), m anti-Oct3/4 (Santa Cruz), rb anti-Oct3/4 (Abcam), rb anti-Sox2 (Abcam), rb anti-cMyc (Santa Cruz), rb anti Klf4 (Abcam), rb anti-Nanog (Millipore), m anti SSEA1 (Thermo Scientific), rb anti-doublecortin x (Abcam), rb anti-Lin28 (Abcam), m anti-alpha smooth muscle actin (Abcam), m anti-alpha feto protein (R&D systems), gt anti-Sox1 (R&D systems), rb anti-Pax6 (Covance), m anti-Tuj1 (Covance), m anti-Ki67 (BD Biosciences). As secondary antibodies Alexa-fluorophore conjugated antibodies (Invitrogen) were used and DNA was counterstained using Hoechst 33258 (Invitrogen). For western blot analysis, the following antibodies were additionally used: m anti-Flag (Sigma), m anti-HA (Roche Applied Sciences), rb anti-GAPDH (Abcam), HRP-linked antibodies (GE Healthcare).
8
Immunohistochemical analysis of mouse brain
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Mice under deep anesthesia were perfused with 50 ml PBS following 50 ml 4% PFA/1 PBS solution. After dissection, isolated brains were post-fixed in 4% PFA/1 PBS solution over night at 4 °C. 40 μm sagittal brain sections were cut using a Vibratom (Leica VT 1200 S). Free-floating sections were permeabilized in Tris-buffered saline solution with 0.1M Tris, 150mM NaCl, pH 7.4/0.5% Triton-X 100/0.1% Na-Azide/0.1% Na-Citrate/5% normal goat serum (TBS+/+/+) for at least 1 h. The primary antibodies anti-Hu Nuclei (1:200; Millipore), anti-DCX (1:400; Abcam), anti-TuJ1 (1:600; Covance) and anti-GFAP (1:100; Millipore) were diluted in TBS+/+/+ and incubated for 48 h on a shaker at 4 °C. For immunofluorescence staining, secondary Alexa-fluorophore conjugated antibodies (Invitrogen) and Hoechst 33258 (1:10000, Invitrogen) were used. Sections were analyzed with a Zeiss LSM 710 confocal microscope.
9
Immunofluorescence Staining Protocol for Tissue Sections
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Free-floating sections were washed 3 times for 10 minutes with TBS with 0.2% Triton X-100 (Roche, Indianapolis, IN) and blocked in 5% Normal Goat Serum (NGS; Jackson ImmunoResearch, West Grove, PA) with 0.2% Triton X-100 in TBS for 1 hour at room temperature. Primary antibodies (see table below) were diluted in 5% NGS in TBS with 0.2% Triton X-100. Sections were incubated overnight at 4°C with primary antibodies and washed three times for 10 minutes with TBS the following morning. Secondary Alexa-fluorophore conjugated antibodies (Invitrogen, Carlsbad, CA) were diluted (1:200) in 5% NGS in TBS with 0.2% Triton X-100, and sections were incubated with secondary antibodies for 2 hours at room temperature, protected from light. After incubation, sections were washed three times for 15 minutes in TBS and mounted with VECTASHIELD with DAPI (Vector Laboratories, Burlingame, CA). Images were acquired on an Olympus FV3000 confocal laser-scanning microscope.
10
Immunofluorescence Labeling Protocol
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Free-floating sections were washed 3 times for 10 minutes with TBS with 0.5% Triton X-100 (Roche, Indianapolis, IN) and blocked in 5% Normal Goat Serum (NGS; Jackson ImmunoResearch, West Grove, PA) with 0.5% Triton X-100 in TBS for 1 hour at room temperature. Primary antibodies (see table) were diluted in 5% NGS in TBS with 0.5% Triton X-100. Sections were incubated overnight at 4°C with primary antibodies and washed three times for 10 minutes with TBS the following day. Secondary Alexa-fluorophore conjugated antibodies (Invitrogen, Carlsbad, CA) were diluted (1:500) in 5% NGS in TBS with 0.5% Triton X-100, and sections were incubated with secondary antibodies for 2 hours at room temperature, protected from light. During the last five minutes of secondary incubation, DAPI was added to achieve a dilution of 1:40,000 (ThermoFisher D1306). After incubation, sections were washed three times for 15 minutes in TBS and mounted with an in-house mounting media (20 mM Tris pH8.0, 90% Glycerol, 0.5% N-propyl gallate). Images were acquired on an Olympus FV3000 confocal laser-scanning microscope.
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