Neurite Outgrowth Reagent Kit

• Neuronal-specific fluorescent reagent identifies neuronal cell bodies and neurites in a heterogeneous cell population without isolation steps
• Validated protocol for sample preparation & processing requires approximately 2.5h post-compound treatment and is automation compatible
• Cells prepared with the HCS Reagent Kit may be analyzed directly via standard fluorescence microscopy; or HCS imaging systems
• No radioactivity, cell lysis, purification, or filtration steps

The Neurite Outgrowth HCS Reagent Kit enables quantification of neurons, neuronal like cells, and neurite outgrowth from a heterogeneous population of live cells growing on standard high-density microplates. The cells can either be neuronal-like cell models, such as PC12 or Neuroscreen™-1 Cells, or primary brain cultures that contain a subpopulation of neurons. The kit includes a primary antibody specific for neuron cell bodies and neurites and DyLight™ 488-conjugated Secondary Antibody. Cell nuclei are identified by DNA-specific Hoechst Dye.

Neurons and neuronal-like cells extend processes known as neurites. A major focus for drug discovery is identifying compounds that affect the growth of neurites. Drugs that promote nerve growth would aid treatment in a wide variety of diseases and traumas that result in neuropathy and nerve injury. Examples of areas that would benefit from neuronal regeneration include spinal cord injuries; neuropathy resulting from diseases such as diabetes; stroke; and neurodegenerative diseases such as Parkinson’s and Alzheimer’s diseases.

The Neurite Outgrowth HCS Reagent Kit is based on immunofluorescence using a validated antibody that specifically labels both neurites and neuronal cell bodies from a wide range of mammalian species. Specific identification of neurons and their neurites from mixed populations of cells such as primary brain cultures is also possible. The Neurite Outgrowth Kit reagents, in combination with the ArrayScan HCS Reader and the (Extended) Neurite Outgrowth BioApplication software enable automated plate handling, focusing, cell image acquisition, analysis, and neurite outgrowth quantification. For a more detailed description of the image processing algorithm, see the Neurite Outgrowth or Extended Neurite Outgrowth BioApplication Guide that accompanies the software.

Figure 1. Application of the Neurite Outgrowth imaging algorithm on a field of PC-12 cells acquired with the 10X objective lens on the ArrayScan® HCS Reader. The left image displays a composite of the nuclear and neuronal raw images acquired by the ArrayScan® HCS Reader.  Cell nuclei (blue) are labeled with Hoechst Dye.  The neurons and neurites (green) are identified by immunofluorescence. The panel on the right shows the neuronal image in grayscale, with the neurite identified by the algorithm overlaid with a green trace.

Figure 2. A primary human brain culture was prepared with the Neurite Outgrowth Kit Reagents. All of the cells nuclei were labeled with Hoechst Dye (left). Only the neurons and their neurites were labeled with the neuron-specific marker (right).

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