Immunofluorescent Staining of FLAG-CASZ1b

Cells were cultured in 8-well Lab-Tek Chamber Slides (Cat. no. 177402) for indicated time period. Cells were fixed, permeabilized, blocked, and stained as described previously^{28 (link)}. For indirect immunofluorescent cell staining, an anti-FLAG M2 monoclonal antibody and an Alexa Fluor 594-conjugated goat anti-mouse antibody were used to detect FLAG-CASZ1b or mutant constructs. An anti-MHC antibody and an Alexa Fluor 594-conjugated goat anti-mouse antibody were used to detect MHC. Stained cells were imaged and analyzed using a Nikon Eclipse TE300 fluorescent microscope.

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Liu Z., Zhang X., Lei H., Lam N., Carter S., Yockey O., Xu M., Mendoza A., Hernandez E.R., Wei J.S., Khan J., Yohe M.E., Shern J.F, & Thiele C.J. (2020). CASZ1 induces skeletal muscle and rhabdomyosarcoma differentiation through a feed-forward loop with MYOD and MYOG. Nature Communications, 11, 911.

Publication 2020

Eclipse TE300 fluorescent microscope

Alexa 594 Anti antibody Cells Goat Indirect immunofluorescent Microscope Mouse

Corresponding Organization : Center for Cancer Research

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Variable analysis

independent variables

FLAG-CASZ1b or mutant constructs

dependent variables

Localization and expression of FLAG-CASZ1b or mutant constructs
Localization and expression of MHC

control variables

Culture conditions (8-well Lab-Tek Chamber Slides)
Fixation, permeabilization, blocking, and staining procedures
Imaging method (Nikon Eclipse TE300 fluorescent microscope)

controls

Positive control: Anti-FLAG M2 monoclonal antibody and Alexa Fluor 594-conjugated goat anti-mouse antibody to detect FLAG-CASZ1b or mutant constructs
Positive control: Anti-MHC antibody and Alexa Fluor 594-conjugated goat anti-mouse antibody to detect MHC

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