MindMap Gallery Detection method of autophagy activity in cell lines
Methods for detecting autophagy activity in cell lines mainly include biochemical assays, flow cytometry, and reporter assays. Biochemical assays assess autophagy activity by measuring the LC3-II/I ratio and p62 protein degradation. Flow cytometry utilizes fluorescently labeled antibodies against autophagy-related proteins for quantitative cellular analysis. Reporter assays utilize GFP-LC3 and mCherry-LC3 fusion protein systems to monitor autophagy flux in real-time.
Edited at 2024-12-24 22:45:02In the introduction section, elastin is a protein that plays a crucial role in the body, endowing tissues with elasticity and toughness. Its properties include high elasticity and good ductility. Elastin has a wide range of sources and is commonly found in animal connective tissues such as skin and blood vessels. When using elastin in products, the amount and method of addition should be determined based on the characteristics of the product. It has many uses and can be added to skincare products to enhance skin elasticity and reduce wrinkles; Improving food texture and enhancing taste in the food industry; In the medical field, it can be used to make elastic scaffolds such as artificial blood vessels, providing support for tissue repair. Reasonable use can play its unique value.
This document aims to guide the proper use of proteins. Proteins are the fundamental substances that make up life and are essential for maintaining normal physiological functions in the body, with a wide variety of types. When using protein, daily diet is an important source, such as meat, eggs, etc., which are rich in high-quality protein. Measuring protein intake can be done using professional tools or methods to ensure that the intake meets the body's needs. When mixing protein powder, it should be prepared according to the recommended ratio, and the water temperature should not be too high to avoid damaging the protein activity. Different groups of people have different protein requirements, such as athletes and fitness enthusiasts who can increase their intake appropriately to meet the needs of body repair and muscle growth.
Introduction to Unsaturated Polyester Resin: It is an important thermosetting resin with various excellent properties. It has a wide range of applications and is used in the construction industry to produce fiberglass products such as doors, windows, decorative panels, etc; Used in the automotive industry for manufacturing body components. The advantages of use include simple molding process, low cost, and chemical corrosion resistance. However, it also faces challenges and limitations, such as relatively poor heat resistance and susceptibility to aging. During use, corresponding protective measures should be taken according to specific application scenarios, such as adding heat-resistant agents, antioxidants, etc., to extend their service life and fully leverage their advantages.
In the introduction section, elastin is a protein that plays a crucial role in the body, endowing tissues with elasticity and toughness. Its properties include high elasticity and good ductility. Elastin has a wide range of sources and is commonly found in animal connective tissues such as skin and blood vessels. When using elastin in products, the amount and method of addition should be determined based on the characteristics of the product. It has many uses and can be added to skincare products to enhance skin elasticity and reduce wrinkles; Improving food texture and enhancing taste in the food industry; In the medical field, it can be used to make elastic scaffolds such as artificial blood vessels, providing support for tissue repair. Reasonable use can play its unique value.
This document aims to guide the proper use of proteins. Proteins are the fundamental substances that make up life and are essential for maintaining normal physiological functions in the body, with a wide variety of types. When using protein, daily diet is an important source, such as meat, eggs, etc., which are rich in high-quality protein. Measuring protein intake can be done using professional tools or methods to ensure that the intake meets the body's needs. When mixing protein powder, it should be prepared according to the recommended ratio, and the water temperature should not be too high to avoid damaging the protein activity. Different groups of people have different protein requirements, such as athletes and fitness enthusiasts who can increase their intake appropriately to meet the needs of body repair and muscle growth.
Introduction to Unsaturated Polyester Resin: It is an important thermosetting resin with various excellent properties. It has a wide range of applications and is used in the construction industry to produce fiberglass products such as doors, windows, decorative panels, etc; Used in the automotive industry for manufacturing body components. The advantages of use include simple molding process, low cost, and chemical corrosion resistance. However, it also faces challenges and limitations, such as relatively poor heat resistance and susceptibility to aging. During use, corresponding protective measures should be taken according to specific application scenarios, such as adding heat-resistant agents, antioxidants, etc., to extend their service life and fully leverage their advantages.
Detection method of autophagy activity in cell lines
Microscopy-based methods
Fluorescence microscopy
Use of GFP-LC3 fusion protein
Autophagosome formation visualized by punctate pattern
Monitoring of autophagic flux
Live-cell imaging
Real-time observation of autophagy
Quantification of autophagosome dynamics
Electron microscopy
Ultrastructural analysis of autophagic compartments
Identification of autophagosomes and autolysosomes
Morphological changes in organelles during autophagy
Biochemical assays
Western blotting
Detection of autophagy-related proteins
LC3 lipidation (LC3-I to LC3-II conversion)
p62/SQSTM1 degradation
Analysis of signaling pathways
mTOR pathway components
AMPK activation
Enzymatic assays
Measurement of lysosomal enzyme activity
Cathepsin B and L activity
β-hexosaminidase activity
Flow cytometry
Analysis of autophagic cells
Detection of GFP-LC3 puncta in cells
Quantification of autophagic cells
Assessment of autophagic flux
Use of fluorescent probes
LysoTracker for acidic compartments
Identification of autolysosomes
Quantification of lysosomal activity
Reporter assays
Luciferase-based assays
Use of autophagy-inducible promoter
Monitoring of autophagy induction
Quantification of promoter activity
Fluorescent protein-based assays
mCherry-GFP-LC3 reporter
Dual color detection of autophagosomes
Differentiation between autophagosomes and autolysosomes
Metabolic labeling
Use of radioactive or non-radioactive amino acids
Pulse-chase experiments
Tracking of protein degradation rates
Measurement of autophagic protein turnover
Stable isotope labeling by amino acids in cell culture (SILAC)
Quantitative proteomics
Relative quantification of protein degradation
Identification of autophagy substrates
Immunofluorescence
Detection of autophagy markers
Co-localization studies
LC3 with lysosomal markers
p62 with autophagosomes
Analysis of subcellular distribution
Visualization of autophagic structures
Quantification of autophagy-related proteins in specific compartments
Genetic approaches
RNA interference (RNAi)
Silencing of autophagy genes
Validation of autophagy-related proteins
Impact on autophagic activity
Overexpression of autophagy regulators
Analysis of autophagy induction or inhibition
Effects on autophagosome formation
Effects on cell survival and death
High-content screening (HCS)
Automated imaging and analysis
Multiparametric readouts
Quantification of autophagosome number and size
Correlation with cell health and viability
Use of fluorescent probes and dyes
Specificity for autophagy detection
LysoTracker, monodansylcadaverine (MDC)
Assay for high-throughput screening of autophagy modulators