During development, neurons extend numerous processes that differentiate into dendrites and axons. These processes (termed neurites) are critical for communication between neurons. Characterization of neurite outgrowth, maturation, and the disruption of neurite networks is key to the study of neuropathological disorders (e.g. neurodegeneration), neuronal injury/regeneration and screening for neurotoxicity.
Measurements of neurite outgrowth are commonly made following antibody labelling or staining of the neuronal cultures. These methods require cell fixing and include multiple, manual wash/preparation steps that ultimately kill the neurons and may disrupt delicate neurite structures. Measurements of neurite length are usually made at a single time point, or at best a limited series of time points, as cultures need to be shuttled into and out of the incubator for analysis. Neurite outgrowth assays are often short term (24 or 48 h) and lack information associated with longer term neuronal biology such as the development and disruption of mature neural networks.
IncuCyte® Neurite Analysis Assays enable automated, continuous analysis of neurite outgrowth and neural network stability - all inside your cell culture incubator. Quantify neurite length and branch points in your choice of neurons (e.g. primary, iPSC-derived, immortalized neuronal cell lines) in monoculture or in co-culture with astrocytes.
Measure changes in neurite length and branch points in mixed cultures of neurons and astrocytes using fluorescent neuronal labelling reagents and IncuCyte analysis software.
Continuous measures of neurite dynamics. IncuCyte® NeuroTrack analysis software automatically analyzes images to provide measures of neurite length, branch points and neuronal cell body clusters.
Measure treatment effects automatically and non-invasively. IncuCyte neurite analysis assays allow every well of a 96- or 384-well plate to be imaged and analyzed automatically to provide a microplate readout of neurite dynamics over time (A). Time-courses reveal concentration-dependent treatment effects (B). Transform data into concentration-response curves to compare pharmacology (C).
Create relevant in vitro models of neurodegeneration and neurotoxicity - screen for neuroprotective treatments. Concentration dependent neurotoxic effects of 6-hydroxydopamine (6-OHDA) measured in human iPSC-derived dopaminergic neurons (Dopa.4U, Axiogenesis) in a functional model of Parkinson’s disease. Treatment with bone derived neural factor (BDNF) promotes neurite formation but provided little or no protection against the effects of 6-OHDA
Neuronal monocultures—label-free protocol overview
Neuronal co-cultures—fluorescence protocol overview
IncuCyte® Cytotox and Annexin V reagents
Use IncuCyte® Cytotox and Annexin V reagents to positively mark and quantify apoptotic or cytotoxic cells. (Left) IncuCyte® Cytotox Red reagent labels dead rat forebrain neurons treated with 333 μM glutamate. (Right) IncuCyte® Annexin V Green reagent identifies apoptotic cells in a rat forebrain neuron and astrocyte co-culture in the presence and absence of 300 μM glutamate. Neurons are selectively labeled with the IncuCyte® NeuroLight Red reagent. Multiplexed measurements of cell viability and neurite dynamics can be made in real time.
Measure time-courses and concentration-dependent effects on neurite dynamics and cell viability. Glutamate excitotoxicity in rat forebrain neurons cultured in the presence of astrocytes: (left) neurite network length is disrupted in a concentration dependent manner, (center) measurements apoptosis reveal early effects on cell health (right) concentration-response curves.
Measure neurite dynamics in monocultures and co-cultures with astrocytes.
|Reagents||IncuCyte® NeuroLight™ Red||0.45 mL||4584|
|IncuCyte® NeuroPrime™ Cell Kit||1 Kit||4585|
|Software||IncuCyte® NeuroTrack™ Software||9600-0010|
Detect and quantify apoptotic cells in real time.
|Reagents||IncuCyte® Annexin V Red for Apoptosis||1 vial, 100 tests||4641|
|IncuCyte® Annexin V Green for Apoptosis||1 vial, 100 tests||4642|
Detect and count non-viable cells in real time.
|Reagents||IncuCyte® Cytotox Red||5 μL x 5||4632|
|IncuCyte® Cytotox Green||5 μL x 5||4633|
Cryopreserved neuronal cells and astrocytes.
|Neuro-2a NucLight™ Green Cells||2 x 106 cells/vial||4511|
|Neuro-2a NucLight™ Red Cells||2 x 106 cells/vial||4512|
|NeuroPrime™ rAstrocytes||2 x 106 cells/vial||4586|
|NeuroPrime™ rForebrain Neurons||2 x 106 cells/vial||4587|