Cells or tissue sections were washed twice in PBS, fixed in freshly prepared 3.7% paraformaldehyde for 30 min at 4°C, washed once in PBS and permeabilized in 0.1% Triton X-100 in blocking solution (3% goat serum plus 0.1% BSA in PBS) for 30 min at room temperature, then washed once in PBS, and left in blocking solution for 2 h. Cells were incubated overnight at 4°C with primary antibodies against Oct4 (sc5279, Santa Cruz), Nanog (ab80892, Abcam), SSEA-1 (MAB4301, Millipore), βIII-Tubulin (CBL412, Chemicon), alpha 1-fetoprotein (AFP; DAK-N1501, Dako), alpha smooth muscle actin (α-SMA; ab5694-100, Abcam) and Nestin (MAB353, Millipore). Then cells were washed three times (each for 15 min) with blocking solution, and incubated for 2 h with secondary antibodies at room temperature. Goat Anti-Mouse IgG (H+L) FITC (115-095-003, Jackson) and Goat Anti-Rabbit IgG (H+L) Alexa Fluor® 594 (111-585-003, Jackson), diluted 1:200 with blocking solution, were used. Samples were washed, and counterstained with 0.5 mg/ml Hoechst 33342 (H1398, MP) in Vectashield mounting medium. Fluorescence was detected and imaged using a Zeiss Axio-Imager Z1 fluorescence microscope.
Goat anti mouse igg h l fitc
Manufactured by Jackson ImmunoResearch
Sourced in Panama
Goat Anti-Mouse IgG (H + L) FITC is a secondary antibody reagent that binds to mouse immunoglobulin (IgG) molecules. The antibody is conjugated to the fluorescent dye FITC (Fluorescein Isothiocyanate), which allows for the detection and visualization of mouse IgG in various immunoassays and applications.
Automatically generated - may contain errors
Lab products found in correlation
Goat anti rabbit igg h l alexa fluor 594, by Jackson ImmunoResearch (4 mentions)
Sc 5279, by Santa Cruz Biotechnology (2 mentions)
Ab80892, by Abcam (2 mentions)
Dak n1501, by Agilent Technologies (2 mentions)
Ab5694 100, by Abcam (2 mentions)
Mab4301, by Merck Group (2 mentions)
Axio imager z1 fluorescence microscope, by Zeiss (2 mentions)
Cbl412, by Merck Group (2 mentions)
Mab353, by Merck Group (1 mentions)
Hoechst33342, by MP Biomedicals (1 mentions)
Ab4340, by Merck Group (1 mentions)
Cd11b fitc, by Miltenyi Biotec (1 mentions)
Lsm 710, by Zeiss (1 mentions)
Fluorsave, by Merck Group (1 mentions)
Bx61 fluorescence microscope, by Olympus (1 mentions)
Fitc goat anti rabbit igg h l, by Jackson ImmunoResearch (1 mentions)
7 protocols using goat anti mouse igg h l fitc
1
Immunofluorescence Analysis of Stem Cell Markers
2
Immunofluorescence Analysis of Stem Cell Markers
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Cells were washed twice in PBS, then fixed in freshly prepared 3.7% paraformaldehyde in PBS (pH 7.4) for 30 min at 4 °C, washed in PBS for one time and permeabilized in 0.1% Triton X-100 in blocking solution (3% goat serum plus 0.1% BSA in PBS) for 30 min at room temperature, then washed in PBS for one time, and left in blocking solution for 2 h. Cells were incubated overnight at 4 °C with primary antibodies against Oct4 (sc5279; Santa Cruz), Nanog (ab80892; Abcam), SSEA-1 (MAB4301; Millipore), βIII-tubulin (CBL412; Chemicon), alpha 1-fetoprotein (AFP; DAK-N1501; Dako), alpha smooth muscle actin (α-SMA; ab5694-100; Abcam), γH2AX (05-636; Millipore), TRF1 (TRF12-S; Alpha Diagnostic) and Zscan4 (AB4340; Millipore). Then cells were washed three times (each for 15 min) with blocking solution, and incubated for 2 h with secondary antibodies at room temperature. Goat Anti-Mouse IgG (H + L) FITC (115-095-003; Jackson) and Goat Anti-Rabbit IgG (H + L) Alexa Fluor® 594 (111-585-003; Jackson), diluted 1:200 with blocking solution, were used. Samples were washed, and counterstained with 0.5 μg/ml Hoechst 33342 (H1398; MP) in Vectashield mounting medium. Fluorescence was detected and imaged using a Zeiss Axio-Imager Z1 fluorescence microscope.
3
Multicolor Flow Cytometry of Immune Cells
4
Immunofluorescent Staining of ESCs
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ESCs were grown on gelatin-treated coverslips and washed twice in PBS, then fixed in freshly prepared 4% paraformaldehyde for overnight at 4 °C, permeabilized in 0.1% Triton X-100 in blocking solution (3% goat serum in PBS) for 30 min, washed three times, and left in blocking solution for 1.5 h. Samples were incubated overnight at 4 °C with primary antibodies (for antibody details, see Supplementary Table S2 ), washed three times, and incubated for two hours with the secondary antibodies, goat anti-mouse IgG (H +L) FITC or goat anti-rabbit IgG (H + L) Alexa Fluor 594 (Jackson). Samples were washed twice with PBS, stained with 0.5 μg/mL DAPI, and mounted in Vectashield mounting medium. Fluorescence was detected and imaged using a Leica Microsystems CMS microscope.
5
Immunofluorescence Staining of Embryonic Stem Cells
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ESCs were grown on gelatin-treated coverslips and fixed with freshly prepared 3.7% paraformaldehyde in PBS, permeabilized in 0.1% Triton X-100 in blocking solution (3% goat serum and 0.1% BSA in PBS) for 30 min, washed three times, and left in blocking solution for 1 h. Samples were incubated overnight at 4 °C with primary antibodies (for antibody details, see Supplementary Table 3 ), washed three times, and incubated for 2 h with secondary antibodies, Goat Anti-Mouse IgG (H + L) FITC or Goat Anti-Rabbit IgG (H + L) Alexa Fluor 594 (Jackson). Samples were washed and counterstained with 0.5 μg/ml Hoechst 33342 or DAPI in Vectashield mounting medium. Fluorescence was detected and imaged using a Zeiss Imager Z1 microscope or Confocal Laser Scanning Microscope (Zeiss LSM710).
6
Immunohistochemical Staining of Brain Sections
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PFA fixed brain sections were quenched (0.1 M glycine, 120 mM phosphate buffer, pH 7.4, 30 min, 4 °C), and permeabilized in blocking buffer (3% Triton X-100; 120 mM phosphate buffer, 1% BSA, pH 7.4, 1 h, 4 °C). Samples were then incubated with primary antibodies (1:200 v/v in blocking buffer; O/N at 4 °C) followed by secondary antibodies (1:200 v/v in blocking buffer, 1–2 h at 23 °C). Samples were washed with blocking buffer (4 °C), rinsed in PBS, and mounted (FluorSave, EMD Millipore). Primary antibodies used were anti-cFOS (rabbit sc-52, SantaCrutz) and anti-NeuN (mouse IgG1 clone A60, EMD Millipore). Secondary antibodies used were: AlexaFluor647 donkey anti-rabbit IgG (H + L) and FITC goat anti-mouse IgG (H + L) (Jackson ImmunoResearch). In some cases, slices were loaded with DAPI (1 μL/mL).
7
Immunofluorescence Analysis of DNA Damage
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Cells were fixed with 3% paraformaldehyde (PFA) for 15 min at room temperature, followed by permeabilization with 0.25% Triton X-100 in phosphate-buffered saline (PBS). Blocking with 1% bovine serum albumin (BSA) in PBS was used prior to immunostaining experiments using the antibodies listed below. Nuclei were visualized by staining with DAPI. The primary antibodies used were γH2AX (1:500, Trevigen, 4418-APC-100) and 53BP1 (1:500 for immunofluorescence, Bethyl, #A300-272A). The secondary antibodies were FITC Goat Anti-Mouse IgG (H + L) (1:500, Jackson ImmunoResearch, 115-095-003) and FITC Goat Anti-Rabbit IgG (H + L) (1:500, Jackson ImmunoResearch, 111-095-144). Images were acquired using the GE IN CELL 2200 high-throughput imaging system, or the Olympus BX61 fluorescence microscope at 40× magnification.
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