Journal: STAR protocols

Article Title: Differentiating PC12 cells to evaluate neurite densities through live-cell imaging.

doi: 10.1016/j.xpro.2022.101993

Figure Lengend Snippet: Figure 1. Schematic of experimental workflow for PC12 cell differentiation

Article Snippet: Optional: If desired, PC12 (NGF-differentiated) lysates (ECM Biosciences Cat#PL7141) can be loaded on the gel as a positive control.

Techniques: Cell Differentiation

Journal: STAR protocols

Article Title: Differentiating PC12 cells to evaluate neurite densities through live-cell imaging.

doi: 10.1016/j.xpro.2022.101993

Figure Lengend Snippet: Figure 2. Critical steps for aliquoting reagents and plating PC12 cells for differentiation (A) Set-up for aliquoting NGF, laminin, and CultureOne in tissue culture hood. Use one or two ice buckets and keep reagent vials and all tubes on ice while aliquoting. (B) Wells to avoid using in a 96-well plate (red line) due to increased potential for evaporation in these wells. (C) Improper (left) and proper (right) technique for plating PC12 cells for differentiation. Cell culture plates should be flat on the hood surface and the pipette should be held vertical when adding cells. Graphics in A and B were created with BioRender.com.

Article Snippet: Optional: If desired, PC12 (NGF-differentiated) lysates (ECM Biosciences Cat#PL7141) can be loaded on the gel as a positive control.

Techniques: Evaporation, Cell Culture, Transferring

Journal: STAR protocols

Article Title: Differentiating PC12 cells to evaluate neurite densities through live-cell imaging.

doi: 10.1016/j.xpro.2022.101993

Figure Lengend Snippet: Figure 3. Different PC12 cell clonal variants vary in their differentiation rates Neurite densities of 4 different PC12 cell clonal variants each containing a different inducible mutant form of the androgen receptor (not expressed in these experiments), were quantified daily during differentiation until the culture reached a neurite density of 1,500 mm/mm2. The time it took for the 4 clonal variants to reach the target neurite density varied from 3–6 days of differentiation, and clonal variants also differed in their propensity to proliferate and clump during differentiation. Representative images of each clonal variant are shown on the day they reached the target neurite density. Three wells per clonal variant were analyzed and data represent mean G SD with a < 0.05. Images are cropped for clarity.

Article Snippet: Optional: If desired, PC12 (NGF-differentiated) lysates (ECM Biosciences Cat#PL7141) can be loaded on the gel as a positive control.

Techniques: Mutagenesis, Variant Assay

Journal: STAR protocols

Article Title: Differentiating PC12 cells to evaluate neurite densities through live-cell imaging.

doi: 10.1016/j.xpro.2022.101993

Figure Lengend Snippet: Figure 4. AraC treatment reduces proliferation and cell clumping in differentiated PC12 cells PC12 cells were differentiated to a neurite density of 1,500 mm/mm2 and treated with 1 mM AraC for 48 h. Three days after AraC treatment, neurite-bearing cells exist as single cells in culture while proliferating cells are greatly reduced. (right, + AraC). In contrast, at the same timepoint in the absence of AraC treatment, neurite-bearing cells exist in clumps (left, - AraC, top) or are overtaken by proliferating cells (left, - AraC, bottom). Images are cropped for clarity.

Article Snippet: Optional: If desired, PC12 (NGF-differentiated) lysates (ECM Biosciences Cat#PL7141) can be loaded on the gel as a positive control.

Techniques:

Journal: STAR protocols

Article Title: Differentiating PC12 cells to evaluate neurite densities through live-cell imaging.

doi: 10.1016/j.xpro.2022.101993

Figure Lengend Snippet: Figure 5. Cell confluency over 6 days of differentiation for western blot analysis Images depicting the confluency of PC12 cells every 2 days of differentiation, with corresponding lysis buffer volume used for cell lysis and average protein concentration in cell lysates. Images are cropped for clarity.

Article Snippet: Optional: If desired, PC12 (NGF-differentiated) lysates (ECM Biosciences Cat#PL7141) can be loaded on the gel as a positive control.

Techniques: Western Blot, Lysis, Protein Concentration

Journal: STAR protocols

Article Title: Differentiating PC12 cells to evaluate neurite densities through live-cell imaging.

doi: 10.1016/j.xpro.2022.101993

Figure Lengend Snippet: Figure 6. NGF-induced neurite outgrowth of PC12 cells over 6 days of differentiation Upon NGF treatment, PC12 cells undergo a morphology change from a circular to triangular-shaped cell body and gradually extend neurites that develop bulbous terminal ends (arrows). Images are cropped for clarity.

Article Snippet: Optional: If desired, PC12 (NGF-differentiated) lysates (ECM Biosciences Cat#PL7141) can be loaded on the gel as a positive control.

Techniques:

Journal: STAR protocols

Article Title: Differentiating PC12 cells to evaluate neurite densities through live-cell imaging.

doi: 10.1016/j.xpro.2022.101993

Figure Lengend Snippet: Figure 7. Differentiated PC12 cells express the neuronal markers Synapsin-1, b-III-Tubulin, and GAP43 (A) Expression of neuronal proteins Synapsin-1, b-III-Tubulin, and GAP43 increase over 6 days of differentiation, corresponding to neurite outgrowth. Phase-contrast images are cropped for clarity. (B) At 6 days of differentiation, neuronal proteins are visualized through immunofluorescence (b-III-Tubulin = red; Synapsin-1 and GAP43 = cyan (pseudo-colored)). Proteins are localized in the nucleus (Synapsin-1 and GAP43), cytoplasm (all), and neurites (all). Arrows indicate expression of Synapsin-1 and GAP43 in the bulbous terminal ends of the neurites. Images are cropped for clarity.

Article Snippet: Optional: If desired, PC12 (NGF-differentiated) lysates (ECM Biosciences Cat#PL7141) can be loaded on the gel as a positive control.

Techniques: Expressing

Journal: STAR protocols

Article Title: Differentiating PC12 cells to evaluate neurite densities through live-cell imaging.

doi: 10.1016/j.xpro.2022.101993

Figure Lengend Snippet: Figure 9. Poor and proper dispersion of PC12 cells in a well An example of poor (left) and proper (right) dispersion of PC12 cells after plating. Cells should be plated as single cells and evenly distributed throughout the well. Images are cropped for clarity.

Article Snippet: Optional: If desired, PC12 (NGF-differentiated) lysates (ECM Biosciences Cat#PL7141) can be loaded on the gel as a positive control.

Techniques: Dispersion