Neurite Outgrowth: Advancing Neurobiology Research with a Powerful Automated Solution

Utilize the full range of supported analysis modes to optimize results and gain deeper insights

Fluorescence-based neurite outgrowth for fixed cell analysis

1. This method supports common immunohistochemistry (IHC) and antibody-based endpoint analysis.
2. The specificity of fluorescence-based analysis is particularly advantageous in mixed and coculture experiments.
3. Leverage multichannel analysis combining neurite/soma antibody signals with nuclear stains for best results (see TekTip 2 for more details).

Single-fluorescence channel analysis for live-cell staining (e.g., Calcein AM)

1. Live-cell fluorescence is facilitated through single-channel fluorescence, allowing for fast and easy labeling of live cells or detection of endogenous fluorescent protein expression.
2. The neurite outgrowth module detects fluorescence signal intensity differences in soma and neurites, allowing accurate segmentation of both compartments for live cell fluorescence-based outgrowth analysis.

Label-free kinetic analysis:

1. Label-free kinetic analysis minimizes sample disruption for long-term kinetic applications while complementing end-point capabilities.
2. Phase contrast imaging generally provides the best results, however standard brightfield imaging methods are also supported.
3. Compatible with the BioSpa 8 automated incubator to support higher throughput kinetic applications.

Achieve more accurate soma boundaries and improved soma counts by employing a nuclear label

There are a couple of notable advantages to neurite outgrowth analysis approaches that include nuclear staining. Labeled nuclei can help resolve and analyze cell bodies that are clustered, resulting in more accurate cell counts and cell segmentation. Additionally, min and max limits in the module can be used to include or discard cells based on size, providing an effective method to exclude dead cells from the analysis due to their relatively condensed size compared to viable cells (Figure 3).

Another advantage of employing nuclear labels is the ability to identify and exclude nuclei lacking sufficient overlap with the antibody signal for neurite compartments. This can help ignore nuclei that are simply positioned near, or underneath, neurite processes emanating from a different cell.

Lastly, using nuclear labels in neurite analysis applications enables the user to finely control the distance the soma mask extends from the nucleus down into the neurite compartment. This feature relies on the “max nucleus/soma distance” setting in the module. This feature is particularly helpful for analyzing immunolabeled samples in which staining in the proximal neurite is typically intense, often equal to soma intensity. Under these conditions, there is often not a clear signal intensity boundary between the soma and neurites. However, the “max nucleus/soma distance” setting enables users to accurately define the soma boundary for different cell types and staining profiles (Figure 4).