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. 2019 Dec;67(6):256-258.
doi: 10.2144/btn-2019-0073. Epub 2019 Oct 17.

Higher percentage of horse serum in culture media blocks attachment of PC12 cells

Jennifer L Meth  1 , Alan R Schoenfeld  1
Affiliations

Higher percentage of horse serum in culture media blocks attachment of PC12 cells

Jennifer L Meth et al. Biotechniques. 2019 Dec.
No abstract available

Keywords: PC12; cell attachment; differentiation; horse serum; neurite outgrowth.

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Conflict of interest statement

Financial & competing interests disclosure

Research reported in this publication was supported by the National Cancer Institute of the National Institutes of Health (award number R15CA121992). The authors have no other relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript apart from those disclosed.

No writing assistance was utilized in the production of this manuscript.

Figures

Figure 1.
Figure 1.. Percent of loosely attached (rounded) PC12 cells.
PC12 cells were plated in standard culture dishes in media containing the indicated combination of horse serum and fetal calf serum. Images captured using a Zeiss Primovert inverted microscope (at 40× magnification) equipped with an Excelis high-definition digital camera after (A) 24 h and (B) 48 h were analyzed blindly, counting the percentage of cells loosely attached, as judged by a halo around the cells in the image and a lack of extensions from the cell body. Cell counts from three images for each combination were averaged. For statistical analyses, ANOVA with Tukey HSD post-hoc tests were performed. (Note: In both (A & B), there was no statistically significant difference between horse serums 1 and 2 at any of the horse serum percentages). (C) A representative image showing loosely attached cells with halos and attached (polygonal) cells.
Figure 2.
Figure 2.. Neurite outgrowth assay.
Wells in six-well plates were incubated with 0.5 ml of a 20 μg/ml solution of laminin (Sigma) and allowed to dry. DMEM media containing the indicated percentages of horse serum along with 5% fetal calf serum were added to the wells (3 wells for each horse serum percentage). PC12 cells (1 × 104 cells) were added to each well and allowed to attach for 48 h. Media was replaced with serum-free DMEM, supplemented with 100 ng/ml of NGF (Becton Dickenson), and the cells were allowed to incubate for another 48 h. Five images from each well were captured using a Zeiss Primovert inverted microscope (at 100× magnification). Cells from each image were counted, both total cells and those with neurites, defined as neuron-like extensions of the cell membrane that were equal to or longer than one diameter of the cell. For statistical analyses, ANOVA with Tukey HSD post-hoc tests were performed.
See this image and copyright information in PMC

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