Lysosome Function Analysis - Selection Guide for Detection and Imaging Reagent / Probe / Kit

Science Note

[July 11, 2023]                                                                                                                                                                                                                            Previous Science Note

Lysosomal pH regulated by LAMP proteins 

Through a combination of various scientific methods, this research reveals the vital role of human lysosome-associated membrane proteins (LAMP-1 and LAMP-2) in maintaining lysosomal pH balance. Despite being mainly recognized as lysosomal markers, LAMP-1 and LAMP-2 directly influence the lysosomal cation channel TMEM175, aiding the acidification of lysosomes to a pH level that optimizes hydrolase activity. Interruptions to the interaction between LAMP and TMEM175 can increase lysosomal pH, impairing their hydrolytic function.
We offer small fluorescent probes for detecting lysosomal mass and pH with several color options: Lysosomal Acidic pH Detection kit-Green/Red and Green/Deep Red. 

Lysosomal LAMP proteins regulate lysosomal pH by direct inhibition of the TMEM175 channel    
Click here for the original article: Jiyuan Zhang, et. al., Mol. Cell. (2023)

Point of Interest
- The lysosomal proteins LAMP-1 and LAMP-2 have a direct interaction with the TMEM175 channel.
- This complex formation between TMEM175 and LAMPs is facilitated by their transmembrane (TM) domains.  
- The binding of LAMP proteins inhibits the channel activity of TMEM175.
- This inhibition of TMEM175 promotes lysosomal acidification, thereby enhancing the efficiency of hydrolase activity.
Related Techniques
           Lipid droplets detection Lysosomal Acidic pH Detection Kit-Green/Red, Green/Deep Red
           Lysosome staining pH-dependent (Red)​ and pH-independent (Green / Deep Red) probes
           Autophagy detection DAPGreen / DAPRed (Autophagosome detection), DALGreen (Autolysosome detection)
           Endocytosis detection ECGreen
           Endocytic internalization assay AcidSensor Labeling Kit 
​​           Plasma membrane staining PlasMem Bright Green / Red
           ​​Total ROS detection Highly sensitive DCFH-DA or Photo-oxidation Resistant DCFH-DA 
 
Related Applications

Accurate Measurement for Lysosomal pH changes

Existing lysosomal pH detection reagents have issues with dye localization, pH sensitivity, and retention. pHLys Green is a dye that solves these issues. The improved dye retention and localization enable detection of normal lysosomes, and the improved pH sensitivity enables detection of slight pH changes.

1. High sensitive pH detection
Comparison of pH response of cells treated with low concentrations of lysosomal acidification inhibitor Bafilomycin A1

2. High specificity for lysosomes
Comparison of specificity for lysosomes using lysosomal marker protein LAMP1-GFP expressing cells

3. High retention in lysosomes
Comparison of intracellular retention

Product in Use:
   - Lysosomal Acidic pH Detection Kit-Green/Deep Red

Related Product:
   - pHLys Red- Lysosomal Acidic pH Detection

 

 

 

Why is Lysosomal Function Important?

The lysosome has been revealed to be an important organelle with a complex role in nutrient sensing and multifaceted signaling. Its importance has been emphasized in research fields as diverse as a neurological disease, cancer, immunity, and senescence.

Recent research reveals that lysosome acidification declines in neurons well before extracellular amyloid deposition, thus lysosomal function is now a hot topic in Alzheimer's disease research.


Lysosomal pH Detection Dye: pHLys Red
pHLys Red has a high sensitivity to lysosomal pH change with more precise localization, which allows slight pH changes to be detected in physiological conditions like the early phase of the disease.

 

 

Lysosomal Mass Detection Dye: LysoPrime Green, Deep Red
Dojindo’s LysoPrime Green / Deep Red more precisely detects lysosomal mass due to their unique specificity for lysosomes, and retention independent of lysosomal pH. In addition, the retention time of the dyes in lysosomes has been extended, allowing data to be acquired over time.

 

Recommended for First Time Users
Lysosomal Acidic pH Detection Kit
Two types of dyes and lysosomal pH acidification inhibitors are available as a set. Positive control is included for immediate testing in your experimental system.


- Lysosomal mass detecting dye: LysoPrime Green
- Lysosomal pH detection dye: pHLys Red
- Lysosomal pH acidification inhibitor: Bafilomycin A1

Application data

 

Endocytic vesicles were labeled by ECGreen and the lysosomal mass and pH were detected separately with LysoPrime Deep Red and pHLys Red. Co-staining with ECGreen and Lysosomal dyes showed the inhibition of endocytic vesicle-fusion induced by Bafilmycin A1.

 

We tried the simultaneous detection of lysosomal and mitochondrial dysfunction in Hela cells treated with CCCP or Antimycin (AN). CCCP and AN are well-known inducers of mitochondrial ROS regarding loss of mitochondrial membrane potential. Recent research showed the result that CCCP induces not only mitochondrial ROS but also lysosomal neutralization. To detect mitochondrial ROS, HeLa cells were labeled by mtSOX Deep Red - Mitochondrial Superoxide Detection, and the lysosomal mass and pH were detected separately with LysoPrime Green and pHLys Red. Co-staining with mtSOX and Lysosomal dyes demonstrated that CCCP causes lysosomal neutralization and mitochondrial ROS induction at the same time.

 

Lysosome Staining Reagents and Kits

Explore Dojindo's wide range of lysosomal staining and pH detection dyes. Choose the following kit or reagent that aligns with your experimental requirements.

Product Name
(Item Code)		 Supported Devices Indicator and Detection Color Dyes and
Fluorescence Properties		 Approximate Number
of Use
Lysosomal Acidic pH Detection Kit-Green/Deep Red (L268) ✓ ✓ ✓ pH pHLys Green
Ex: 488 nm / Em: 490-550 nm		 [for 1 set]
35 mm dish: 10 dishes
μ-Slide 8 well: 10 plates
96-well Plate: 2 plates
quantity LysoPrime Deep Red
Ex: 633 nm / Em: 640-700 nm
Lysosomal Acidic pH Detection Kit-Green/Red (L266) ✓ Need G/Y Laser
G:532 nm
Y:561 nm		 ✓ pH pHLys Red
Ex: 561 nm / Em: 560-650 nm
quantity LysoPrime Green
Ex: 488 nm / Em: 500-600 nm
pHLys Red- Lysosomal Acidic pH Detection  (L265) ✓ ✓ pH pHLys Red
Ex: 561 nm / Em: 560-650 nm		 [for 1 tube]
35 mm dish: 10 dishes
μ-Slide 8 well: 10 plates
96-well Plate: 2 plates
LysoPrime Deep Red - High Specificity and pH Resistance  (L264) ✓ ✓ ✓ quantity LysoPrime Deep Red
Ex: 633 nm / Em: 640-700 nm
LysoPrime Green- High Specificity and pH Resistance  (L261) ✓ ✓ ✓ quantity LysoPrime Green
Ex: 488 nm / Em: 500-600 nm		 [for 10 μl]
35 mm dish: 10 dishes
μ-Slide 8 well: 10 plates
96-well Plate: 2 plates

Product Classification

Product Classification

Search word