Necrosis Caco-2

A key signature for necrotic cells is the permeabilization of the plasma membrane. Necrosis can be quantified by several cellular and biochemical assays. When studied minutely, it reveals the difficulty in confirmation in secondary induction of necrosis in apoptotic cells. Apoptotic cells are being analyzed to shift to necrotic status owing to membrane permeability at later stages, and thus, discrimination of two cell death becomes critical. Therefore, it is crucial to use a necrosis detection kit or a defined procedure to analyze this unprogrammed form of death in response to immense chemical and physical insults.

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Found 2 matching solutions for this experiment

Acridine Orange solution

Sigma-Aldrich

Sigma-Aldrich

Upstream tips	Protocol tips	Downstream tips
• Dual Staining Procedure for differentiating Live/ Apoptotoic/Necrotic cells	• Seed 3x10^4 cells onto a 10mm coverslip and incubate overnight to form a confluent monolayer	• The operator to be blinded to the experimental groups and random fields to be selected (40X objective). • A total of 300 attached cells per cover-slip can be morphologically identified and counted as being either necrotic (red/orange nuclei),apoptotic (green condensed or fragmented nuclei) green or live (green non-condensed ovoid or rounded nuclei)

Upstream tips
• Dual Staining Procedure for differentiating Live/ Apoptotoic/Necrotic cells
Protocol tips
• Seed 3x10^4 cells onto a 10mm coverslip and incubate overnight to form a confluent monolayer
Downstream tips
• The operator to be blinded to the experimental groups and random fields to be selected (40X objective). • A total of 300 attached cells per cover-slip can be morphologically identified and counted as being either necrotic (red/orange nuclei),apoptotic (green condensed or fragmented nuclei) green or live (green non-condensed ovoid or rounded nuclei)

Publication protocol 1 Relevant paper

Apoptosis/ Necrosis Assay Kit (blue, green, red) (ab176749)

Abcam

Upstream tips	Protocol tips	Downstream tips
• Apoptosis/ Necrosis Detection Kit (blue, green, red) (ab176749) is designed to simultaneously monitor apoptotic, necrotic and healthy cells. • The PS sensor used in this kit has green fluorescence (Ex/Em = 490/525 nm) upon binding to membrane PS. • In addition, this kit also provides a live cell cytoplasm labeling dye, CytoCalcein Violet 450 (Ex/Em = 405/450 nm), for labeling living cell cytoplasm.	• Seed 2.5X10^4 cells/well in 24-well plates

Upstream tips
• Apoptosis/ Necrosis Detection Kit (blue, green, red) (ab176749) is designed to simultaneously monitor apoptotic, necrotic and healthy cells. • The PS sensor used in this kit has green fluorescence (Ex/Em = 490/525 nm) upon binding to membrane PS. • In addition, this kit also provides a live cell cytoplasm labeling dye, CytoCalcein Violet 450 (Ex/Em = 405/450 nm), for labeling living cell cytoplasm.
Protocol tips
• Seed 2.5X10^4 cells/well in 24-well plates

Manufacturer protocol 1 Relevant paper

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