For the immunofluorescence assay, PLC-PRF-5 and Hep3B cells subjected to different treatments were washed three times with 1 × PBS, fixed in 4% paraformaldehyde (pre-cooled at 4 °C, Solarbio) for 20 min, and blocked with 5% bovine serum albumin (BSA, KeyGEN BioTECH) containing 0.1% TritonX-100 (Sigma) for 30 min at room temperature. Then, the resultants were incubated with anti-E-cadherin (CST, 14472, 1:50) and anti-Vimentin (CST, 5741, 1:100) or anti-SLUG (Affinity, DF6202, 1:100) and anti-USP5 antibodies (Abcam, ab154170, 1:100). After washing with 1 × PBS again, and then incubated with fluorescently conjugated secondary antibodies (1:200, KeyGEN BioTECH) diluted in 5% BSA for about 50 min at room temperature. Finally, the cells were washed with 1 × PBS and mounted with the DAPI-containing mounting medium (Solarbio). The images of cells were taken with a laser scanning confocal microscope (Leica).
Fluorescently conjugated secondary antibodies
Manufactured by Keygen Biotech
Sourced in China
Fluorescently conjugated secondary antibodies are laboratory reagents used to detect and visualize target proteins or biomolecules in various analytical techniques. These antibodies are designed to bind to the primary antibodies that are specific to the target of interest, thereby enabling the detection and localization of the target through the fluorescent signal.
Automatically generated - may contain errors
Lab products found in correlation
Dapi containing mounting medium, by Solarbio (2 mentions)
Triton x 100, by Merck Group (2 mentions)
Bovine serum albumin bsa, by Keygen Biotech (2 mentions)
Paraformaldehyde (pfa), by Solarbio (2 mentions)
Ab154170, by Abcam (1 mentions)
Confocal laser scanning microscope, by Leica camera (1 mentions)
Df6202, by Abcam (1 mentions)
Anti stat3, by Affinity Biosciences (1 mentions)
Anti e cadherin, by Affinity Biosciences (1 mentions)
Anti vimentin, by Affinity Biosciences (1 mentions)
Laser scanning confocal microscope, by Nikon (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Solarbio (1 mentions)
3 protocols using fluorescently conjugated secondary antibodies
1
Immunofluorescence Assay for Cell Markers
2
Immunofluorescence Analysis of Cell Markers
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The PLC/PRF/5 and HepG2 cells with different treatments were washed three times with 1× PBS, fixed in 4% paraformaldehyde (precooled at 4 °C, Solarbio) for 20 min, and blocked with 5% bovine serum albumin (BSA, KeyGen Biotech) containing 0.1% Triton X-100 (Sigma) for 30 min at room temperature. Then, the resultants were incubated with anti-E-cadherin, anti-vimentin, and anti-STAT3 (1:100, Affinity). The cells were washed with 1× PBS again and subsequently incubated with fluorescently conjugated secondary antibodies (1:200, KeyGen Biotech) diluted in 5% BSA for approximately 50 min at room temperature. Finally, the cells were washed with 1× PBS and mounted with the DAPI-containing mounting medium (Solarbio). Images were obtained with a laser scanning confocal microscope (Nikon, Japan).
3
Immunofluorescence Assay for OVCAR-3 Cells
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OVCAR-3 cells cultured on climbing films in 24-well plate were treated with Selene and SeNPs. Then, the cells were fixed in 4% paraformaldehyde for 15 min at room temperature and incubated in 0.01% Triton X-100 solution for 15 min. Subsequently, the cells were blocked with 5% BSA for 1.5 h. The primary antibody diluted with 1% BSA (1:200) was added in the cells, which were then incubated at 4 °C overnight. After the cells were washed with PBS, they were incubated with fluorescently conjugated secondary antibodies (1:200, KeyGen Biotech) for 1 h. The samples were counterstained with mounting medium containing DAPI (Solarbio, China) for 2 min. Finally, the cells were observed under a confocal microscope (Nikon, Japan).
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