Journal: STAR Protocols

Article Title: Protocol for cytoskeleton staining of the semi-adherent multiple myeloma cell line RPMI 8226 by immunofluorescence

doi: 10.1016/j.xpro.2024.103060

Figure Lengend Snippet: Immunofluorescence staining of RPMI 8226 trRpn13-MM2 cells Z-slices from a confocal microscopy image of a sample prepared following the described method showing RPMI 8226 trRpn13-MM2 cells undergoing mitosis (left) and at interphase (right) with fluorescence staining for F-actin (phalloidin-ATTO643, red), β-tubulin (TUBB4A-CF568, yellow) and DNA (DAPI, blue). Membrane blebbing is indicated by a white arrow. Scale bar is 2 μm.

Article Snippet: We immunostained the RPMI 8226 cells with TUBB4A antibodies at a final concentration of 10 μg /mL from 1:30 dilution of a 300 μg /mL stock concentration obtained following the probe conjugation with Biotium CF568 Mix-n-Stain kit according to the manufacturer’s instructions: https://biotium.com/wp-content/uploads/2017/10/PI-Mix-n-Stain-Antibody-Labeling-Kits.pdf . c. Cover the Eppendorf tubes with aluminum foil to prevent light exposure and incubate them horizontally ∼15 h on a rocking platform (VWR Model 100 at the setting 2) at 4°C.

Techniques: Immunofluorescence, Staining, Confocal Microscopy, Fluorescence, Membrane