Cell Autophagy Imaging Analysis

Cell Autophagy Imaging Analysis

Autophagy is an orderly circulatory system within a cell that can degrade unnecessary cytoplasmic proteins and damaged organelles. It is a dynamic autophagy process that exists in all eukaryotic cells. Autophagy is important for cells to resist intracellular and extracellular stress and maintain metabolic homeostasis. It is involved in many physiological events, including cell renewal, cell growth, immunity and aging. Recent evidence indicates that changes in autophagy often occur in many human diseases, such as neurodegenerative diseases, cancer, and cardiomyopathy. Therefore, finding new drugs that target the autophagy pathway is essential for the treatment of human diseases.

Mechanism of cellular autophagy.Figure 1. Mechanism of cellular autophagy.

Autophagy Imaging Analysis

Currently, autophagy detection methods include fluorescence microscopy, biochemical measurement, and western blotting, but these methods are time-consuming, labor-intensive, and require a lot of experience to accurately explain. Image cytometry can generate fluorescence intensity data comparable to flow cytometry and is less time-consuming. It has been widely used in indicating the level of autophagy activity in living cells.

Analysis method Immunofluorescence microscopy Western blot Flow cytometry Transmission electron microscopy Image cytometer
Result provided Area or foci of LC3B per cell Specimen average LC3B expression Average intensity of LC3B or acridine orange per cell Morphological structures of autophagosomes and autolysosomes in cells Average intensity of acridine orange per cell
Sensitivity for basal autophagy + + + + + + + + + + + + +
Sensitivity for autophagic flux + + + + + + + + + + + + + + + +
Assay time ∼8 h ∼6 h ∼4+ h ∼10+ h ∼30–45 min
Sample prep. cost + + + + + + + + + + +
Instrument cost + + + + + + + + + + + + +
Quantification reliability + + + + + + + + + + + + +

Autophagy Imaging Analysis Workflow

The following is the workflow of cell imaging to detect autophagy.

Step 1

Plate cells

Plate adherent or suspended cells into 96 or 384 well plates. Incubate the cells under normal culture conditions.

Step 2

Treat cells

If necessary in the experimental procedure, the cells were treated with various concentrations of the target compound.

Step 3

Marker stain

The treated cells are stained with fluorescent dyes to track autophagosomes.

Step 4

Acquire images

Move the target cell sample into a disposable counting chamber to capture fluorescence and brightfield images.

Step 5

Statistical Analysis

Use the cell imaging analysis software to analyze the autophagy.

Delivery

Average fluorescence intensity
EC50 values
Total number of cells
Other relevant data

Our Advantages

  • Convenient and rapid image acquisition and analysis
  • Rapid detection of autophagy in cell populations
  • High accuracy rate, reducing data discrepancies caused by operations
  • Experienced scientists provide experimental consultation
  • Reduced workload and cost

CD BioSciences can provide you with high-quality and low-cost autophagy imaging and analysis services. We use image cytometry to detect fluorescently labeled autophagosomes, and then quickly evaluate the autophagy response of living cells when treated with various chemical or biological reagents. If you have any needs for autophagy analysis, please feel free to contact us.

Reference
  1. SenthilKumar G, Skiba J H, Kimple R J. High-throughput quantitative detection of basal autophagy and autophagic flux using image cytometry[J]. Biotechniques, 2019, 67(2): 70-73.

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Please note: Our services can only be used for research purposes. Do not use in diagnostic or therapeutic procedures!

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