IF experiments were performed as previously described (Welshhans and Bassell, 2011 (link)), and the following primary antibodies were used: mouse anti-paxillin antibody (1:500), mouse anti-vinculin antibody (1:500), rabbit anti-RACK1 antibody (1:500; Santa Cruz Biotechnology), mouse anti-RACK1 antibody (1:500; Santa Cruz Biotechnology), rabbit anti-ribosomal protein S6 antibody (1:100; Santa Cruz Biotechnology), mouse anti-ribosomal protein S6 antibody (1:100; Santa Cruz Biotechnology), and mouse anti-ribosomal RNA 5.8s Y10b antibody (1:100, Abcam). The following secondary antibodies were used: goat anti-mouse Alexa 488, goat anti-rabbit Alexa 488, goat anti-mouse Cy3 and donkey anti-mouse Alexa 647 (all from Jackson ImmunoResearch). Rhodamine phalloidin or Alexa Fluor 350 phalloidin (Life Technologies) was also added in experiments during the secondary antibody incubation to stain F-actin for visualization of growth cone area.
Protocol detail
Immunocytochemistry for Cellular Components
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mouse anti-vinculin antibody
goat anti-mouse Alexa 488
goat anti-rabbit Alexa 488
goat anti-mouse Cy3
donkey anti-mouse Alexa 647
Rhodamine phalloidin
Alexa Fluor 350 phalloidin
Alexa fluor 350
Anti antibody
Antibodies
Antibody
Donkey
F actin
Goat
Growth cone
Mouse
Mouse ribosomal protein s6
Paxillin
Phalloidin
Rabbit
Rhodamine phalloidin
Ribosomal protein s6
Ribosomal rna
Stain
Vinculin
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