~103 SK-Hep1 and Hep 3B cells were grown on cover glass in 24-well plate to ~70% confluence. The cells were washed with PBS 1X and incubated with 4 μM of either target or control peptide for 3 min. The cells were then washed 3X in PBS, fixed with 4% paraformaldehyde (PFA) for 8 min, washed 3X with PBS then incubated with 2% BSA, 1% goat serum in PBS for 30 min. The cells were incubated with a 1:3000 dilution of primary recombinant rabbit anti-CD44 antibody (#ab189524, Abcam) for 30 min on ice and then incubated with a 1:500 dilution of AF488-labeled secondary goat ant-rabbit immunoglobulin G antibody (#A-11029, Life Technologies) for 12 h at 4 °C, and then mounted on glass slides with ProLong Gold reagent containing DAPI (Invitrogen). Confocal fluorescence images were collected on Leica SP8 confocal microscope using a 63X oil-immersion objective. Fluorescence intensities were quantified using custom MATLAB (Mathworks) software.
Af488 labeled secondary goat ant rabbit immunoglobulin g antibody
Manufactured by Thermo Fisher Scientific
The AF488-labeled secondary goat anti-rabbit immunoglobulin G (IgG) antibody is a fluorescently-labeled antibody used in immunoassays and other applications that require the detection of rabbit primary antibodies. The AF488 fluorophore provides green fluorescence when excited at the appropriate wavelength.
Automatically generated - may contain errors
Lab products found in correlation
Prolong gold reagent, by Thermo Fisher Scientific (3 mentions)
Matlab, by MathWorks (1 mentions)
Sp8 confocal microscope, by Leica (1 mentions)
Sp5 inverted 2 photon flim confocal, by Leica (1 mentions)
Primary monoclonal rabbit anti cmet antibody, by Cell Signaling Technology (1 mentions)
Matlab software, by MathWorks (1 mentions)
2 protocols using af488 labeled secondary goat ant rabbit immunoglobulin g antibody
1
CD44 Expression Quantification in Liver Cancer Cells
2
Quantitative Analysis of cMet Expression
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The cells were blocked with 1X PBS plus 2% BSA for 1 hour at 4 °C, and were incubated with 5 μM of peptide for 10 min at room temperature (RT) in the dark, washed 3×, and fixed with 4% PFA for 5 min, washed with 1X PBS, and then mounted on glass slides with ProLong Gold reagent containing DAPI (Invitrogen). A 1:3000 dilution of primary monoclonal rabbit anti-cMet antibody (#8198, Cell Signaling Technology) was incubated with the cells in vitro as a positive control. Afterwards, the cells were incubated with a 1:500 dilution of AF488-labeled secondary goat ant-rabbit immunoglobulin G antibody (#A-11029, Life Technologies), and then mounted on glass slides with ProLong Gold reagent containing DAPI. Confocal fluorescence images were collected using a 63X oil-immersion objective (Leica SP5 Inverted 2-Photon FLIM confocal). Fluorescence intensities were quantified using custom Matlab software (Mathworks).
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