Neuronal Immunofluorescence Staining

Neurons were fixed in 4% paraformaldehyde/4% sucrose solution for 15 min at 37 °C, permeabilized with 0.25% Triton X-100, and blocked with 0.5% fish skin gelatin at 7 and 16 DIV^{26 (link)}. The following antibodies were used: anti-SMI312 (BioLegend 837904, mouse IgG1/IgM, 0.5 μg mL⁻¹), anti-MAP2 [Abcam ab5392, chicken polyclonal IgY (IgG), 2.5 μg mL⁻¹], Alexa 488-labeled goat anti-mouse IgG1 (Molecular Probes A21121, 2 μg mL⁻¹), and Alexa 568-labeled goat anti-chicken IgG (Molecular Probes A11041, 8 μg mL⁻¹).

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Matsumura R., Yamamoto H., Hayakawa T., Katsurabayashi S., Niwano M, & Hirano-Iwata A. (2018). Dependence and Homeostasis of Membrane Impedance on Cell Morphology in Cultured Hippocampal Neurons. Scientific Reports, 8, 9905.

Publication 2018

anti-SMI312 ab5392 Alexa 488-labeled goat anti-mouse IgG1 A21121

Alexa 568 Anti igg Antibodies Chicken Fish skin Gelatin Goat Igg1 Map2 Molecular probes Mouse Neurons Paraformaldehyde Sucrose Triton x 100

Corresponding Organization : Tohoku University

Other organizations : Fukuoka University

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Variable analysis

independent variables

Fixation duration (15 min)
Fixation temperature (37 °C)
Permeabilization with 0.25% Triton X-100
Blocking with 0.5% fish skin gelatin

dependent variables

Expression of SMI312 (neurofilament marker)
Expression of MAP2 (microtubule-associated protein 2)

control variables

Neuronal culture at 7 and 16 DIV

controls

Positive control: Neurons stained with SMI312 and MAP2 antibodies
Negative control: Neurons stained without primary antibodies

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