Reporter gene assay

A gross majority of classical apoptotic attributes can be quantitatively examined by flow cytometry, the preferred platform for rapid assessment of multiple cellular attributes at a single-cell level. However, sample preparation for such flow cytometry-based techniques could be challenging. Cell harvesting by trypsinization, mechanical or enzymatic cell disaggregation from tissues, extensive centrifugation steps, may all lead to preferential loss of apoptotic cells. To overcome this strictly follow manufacturers instruction of the detection kit.

Get tips on using RealTime-Glo™ Annexin V Apoptosis and Necrosis Assay to perform Necrosis LNCaP

Get tips on using RealTime-Glo™ Annexin V Apoptosis and Necrosis Assay to perform Necrosis HUVEC

Get tips on using RealTime-Glo™ Annexin V Apoptosis and Necrosis Assay to perform Necrosis A549

Get tips on using RealTime-Glo™ Annexin V Apoptosis and Necrosis Assay to perform Necrosis HCT 116

As autophagy is a multi-step process which includes not just the formation of autophagosomes, but most importantly, flux through the entire system, including the degradation upon fusion with lysosomes, which makes it quite challenging for detection. There are several methods for detection in mammalian cells, including immunoblotting analysis of LC3 and p62 and detection of autophagosome formation/maturation by fluorescence microscopy, Currently, there is no single “gold standard” for determining the autophagic activity that is applicable in every experimental context, hence it is recommended to go for the combined use of multiple methods to accurately assess the autophagic activity in any given biological setting.