Reliable Lysosome Visualisation Under Real Experimental Conditions
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Lysosomes sit at the heart of cellular homeostasis, acting as key regulators of degradation, recycling, and signalling pathways. However, accurately monitoring lysosomal function remains a persistent challenge for researchers. Their activity depends on tightly controlled parameters, acidic pH, active hydrolases, and dynamic crosstalk with other organelles, making functional detection inherently complex. Many conventional lysosomal probes fall short where it matters most. Signal loss under acidic conditions, poor specificity, and limited retention often compromise experimental outcomes, leading to incomplete or misleading data. LysoPrime™ Green and LysoPrime™ Deep Red, developed by Dojindo, are designed to address these limitations directly, enabling robust, reliable lysosome visualisation even in demanding experimental settings. |
Why Choose LysoPrime™?
Unlike traditional dyes, LysoPrime probes are engineered with the end user in mind: delivering consistency, accuracy, and confidence in your lysosomal assays.
As illustrated in Figure 1, LysoPrime probes overcome the key limitations of conventional dyes, enabling more faithful monitoring of lysosomal function in living cells.
Figure 1: Graphical scheme showing the differences between conventional dyes vs LysoPrime to monitor lysosome function in living cells.
Key Benefits Aligned with Your Research Needs
1. High Specificity for Lysosomes
Precise localisation is critical when studying lysosomal dynamics. LysoPrime probes demonstrate strong co-localisation with established lysosomal markers, ensuring that your signal reflects true lysosomal structures rather than off-target staining.
As demonstrated in Figure 2, LysoPrime Deep Red shows excellent overlap with LAMP1-GFP in HeLa cells, providing confidence in your imaging data.
What this means for you:
- Reduced background noise
- More accurate interpretation of lysosomal behaviour
- Increased reproducibility across experiments
Figure 2: Comparison between LysoPrime Deep Red and a competitor dye on lysosomal localisation using HeLa cells expressing LAMP1-GFP. (White indicates co-localisation).
Figure 3: LysoPrime Green retention in lysosomes following pH disruption.
Figure 4: Comparative performance of LysoPrime Green and LysoPrime Deep Red.
2. Robust pH Resistance
A major limitation of conventional probes is their dependence on acidic pH for fluorescence. When lysosomal pH fluctuates, whether due to experimental treatment or biological processes, signal integrity is often lost.
LysoPrime probes are different.
Both LysoPrime Green and Deep Red remain localised and fluorescent even when lysosomal acidity is disrupted. As shown in Figure 3, treatment with Bafilomycin A1 does not compromise signal retention.
What this means for you:
- Reliable readouts during pH-modulating experiments
- Compatibility with drug screening and perturbation studies
- Confidence in longitudinal analyses
3. High Retention for Long-Term Studies
Time-course experiments require probes that remain stable within lysosomes. Many dyes rapidly diffuse or degrade, limiting their usefulness for extended observations.
LysoPrime probes exhibit exceptional retention, remaining within lysosomes even after prolonged incubation.
As shown in Figure 5, LysoPrime Deep Red outperforms competitor dyes after 24 hours of staining.
What this means for you:
- Fewer re-staining steps
- Reduced variability over time
- Enhanced suitability for live-cell imaging and long-term assays
Figure 5: Comparison between LysoPrime Deep Red and competitor dyes after 24 hours.
Designed for Complex Lysosome Biology
Supporting Your Research Beyond Detection
Whether you are investigating lysosomal dysfunction in disease, assessing drug-induced perturbations, or exploring autophagy pathways, LysoPrime probes provide the robustness and precision required for advanced research.
They are particularly well-suited for:
- Live-cell imaging under dynamic conditions
- Studies involving lysosomal stress or pH modulation
- Longitudinal and high-content screening assays
Lysosomes are increasingly recognised as central players in a wide range of pathologies, from neurodegeneration to cancer.
To explore this topic further, see our dedicated resource:
“Lysosomes at the Crossroads of Health and Disease: Solutions for In-Depth Functional Studies”
For a quotation or further information, please do not hesitate to contact us.
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Reference
Article content created by Tebubio using courtesy materials provided by Dojindo Europe.
Technical FAQ
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Are there any advices when observing the senescent cells?
Lipofuscin is a fluorescent pigment that accumulates in a variety of cell types with age. Lipofuscin consists of autofluorescent granules and may results in high background for fluorescence microscopy. In order to achieve accurate SA-β-gal activity assay in senescent cells, we recommend to prepare samples without SPiDER-βGal staining. Please compare fluorescence intensity of both cells with or without SPiDER-βGal staining.
For Flow Cytometry Detection
Step 1. Prepare senescent cells and non-senescent cells. Measure MFI (Mean Fluorescence Intensity) of samples below.
Senescent cells
Sample A: The cells stained with SPiDER-βGal
Sample B: The cells without SPiDER-βGal stainingNon-senescent cells
Sample A’: The cells stained with SPiDER-βGal
Sample B’: The cells without SPiDER-βGal stainingStep 2. Calculate SA-β-gal activity (senescent cells) with the following formula
SA-β-gal activity (senescent cells) = MFI of Sample A - MFI of Sample BStep 3. Calculate SA-β-gal activity (non-senescent cells) with the following formula:
SA-β-gal activity (non-senescent cells) = MFI of Sample A’ - MFI of Sample B’Determine the SA-β-gal activity by comparing the SA-β-gal activity between senescent cells and non-senescent cells.
Change of SA-β-gal activity associated with senescence = (Value from Step 2- value from Step 3)For Microscopy
Step 1. Prepare senescent cells without SPiDER-βGal staining and observe fluorescent image.
Step 2. Adjust detection sensitivity in microscopy to reduce background autofluorescence of lipofuscin.
Step 3. Observe fluorescent image of senescent cells and non-senescent cells under the settled condition in step 2. -
What if I see background in control cells?
Check if reagents were stored correctly. SPiDER-βGal DMSO stock solution and Bafilomycin A1 DMSO stock solution are stable for 1 month at -20 ℃. SPiDER-βGal working solution and Bafilomycin A1 working solution can’t be stored. Be sure to use the working solution immediately.
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What if I see background in senescent cells without adding SPiDER-βGal working solution?
Lipofuscin, which consists of autofluorescent granules, accumulated in the senescent cells. In order to achieve accurate SA-β-gal activity assay in senescent cells, we recommend preparing samples without SPiDER-βGal staining. Please compare fluorescence intensity of both cells with / without SPiDER-βGal staining.
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What if I see background in fixed cells assay?
The incubation with SPiDER-βGal working solution was done in a 5% CO2 incubator. We recommend not to use a 5% CO2 incubator during incubation with SPiDER-βGal working solution. If incubation is done in a 5% CO2 incubator, the pH of the buffer may become acidic. Acidic pH results in higher background from the endogenous β-galactosidase activity and it would be difficult to distinguish between control cells and senescent cells. Please incubate the plate in a dry incubator without CO2.
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No difference in fluorescence intensity between senescent cells and control cells
Cellular senescence wasn’t induced. Please make sure to prepare a positive control.
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Low fluorescence Reading
• Incubation time with SPiDER-βGal working solution was short. Optimize the incubation time (～ 60 min).
• Concentration of SPiDER-βGal working solution was low. Test different ranges. -
Background in fix cells assay
We recommend not to use a 5% CO2 incubator during incubation with SPiDER-βGal working solution. If incubation is done in a 5% CO2 incubator, the pH of the buffer may become acidic. Acidic pH results in higher background from the endogenous β-galactosidase activity and it would be difficult to distinguish between control cells and senescent cells. Please incubate the plate in a dry incubator without CO2.
Take Control of Your Lysosomal Imaging
With LysoPrime Green and LysoPrime Deep Red, you gain more than just a dye; you gain a reliable tool tailored to the realities of your experiments. Achieve clearer insights. Reduce uncertainty. Advance your research with confidence.