HEK293T cells were transfected with plasmids encoding mouse Juno (clone B2 from the cDNA library) or with GFP-tagged full-length mouse Izumo1 (clone MG222708, Origene, USA) using Lipofectamine 2000 (Invitrogen) according to the manufacturer’s protocol. The day after transfection, equal numbers of cells were mixed and examined for binding and fusion using confocal microscopy two and 24 hours later. Izumo1 was detected by the GFP tag and Juno using the anti-Juno antibody by immunocytochemistry. Briefly, cells were incubated with anti- Juno antibody, diluted 1:200 in PBS/2 % BSA, washed in PBS followed by anti-rat Alexa Fluor 568-conjugated secondary antibody (Molecular Probes). Finally, nuclei were stained with DAPI (Slow Fade mounting medium, Molecular Probes).
Anti rat alexa fluor 568 conjugated secondary antibody
Manufactured by Thermo Fisher Scientific
The Anti-rat Alexa Fluor 568-conjugated secondary antibody is a fluorescently labeled antibody designed to detect rat primary antibodies in immunological applications. The Alexa Fluor 568 dye provides a bright, photostable fluorescent signal that can be detected using standard red fluorescence microscopy and flow cytometry techniques.
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2 protocols using anti rat alexa fluor 568 conjugated secondary antibody
1
Juno and Izumo1 Mediate Cell Fusion
2
Juno and Izumo1 Mediate Cell Fusion
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HEK293T cells were transfected with plasmids encoding mouse Juno (clone B2 from the cDNA library) or with GFP-tagged full-length mouse Izumo1 (clone MG222708, Origene, USA) using Lipofectamine 2000 (Invitrogen) according to the manufacturer’s protocol. The day after transfection, equal numbers of cells were mixed and examined for binding and fusion using confocal microscopy two and 24 hours later. Izumo1 was detected by the GFP tag and Juno using the anti-Juno antibody by immunocytochemistry. Briefly, cells were incubated with anti- Juno antibody, diluted 1:200 in PBS/2 % BSA, washed in PBS followed by anti-rat Alexa Fluor 568-conjugated secondary antibody (Molecular Probes). Finally, nuclei were stained with DAPI (Slow Fade mounting medium, Molecular Probes).
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