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SpheroCHECK SPAchip® pH Green Single-Detection Kit
SpheroCHECK SPAchip® pH Green Single-Detection Kit
Ref. SS-001-PHG
SpheroCHECK SPAchip® pH Green Single-Detection Kit can be used on spheroids and 3D-biology to measure intracellular and extracellular ph levels by changes in fluorescence intensity, which allows a more comprehensive study of spheroid’s establishment, metabolism, and viability.
SpheroCHECK SPAchip® pH Green Single-Detection Kit enables continuous, simultaneous, and accurate monitoring of intracellular and extracellular pH in living 3D models such as spheroids. Evaluation of pH levels is key for effective quality control of living cells to warrant the standardization and reproducibility of your cell experimental methodologies and enable a more comprehensive study of the spheroid formation and its physiology.
SpheroCHECK SPAchip® assay kits are novel cell-based assays for living single-cell developed by A4cell that bring together the fields of nanotechnology and cell biology. SpheroCHECK SPAchip® kits are composed of fluorescently labeled silicon microparticles -SPAchip®- that can be internalized in cultured spheroids to monitor changes in specific intracellular analyte concentrations for long periods of time. Fluorescence intensity is detected from individual SPAchips harboring a pH-sensitive chemical probe covalently attached to their surface.
SpheroCHECK SPAchip® Single-Detection kits are optimized for its use with confocal microscopes and HCS/HCA analyzers with 20X or over magnification objectives, yet widefield fluorescence microscopes and imaging system with fixed wavelength filters can be also used. Flow cytometers have additionally been validated to analyze intra and extracellular SPAchips.
SPAchip® kits can be handled as common fluorophores and chemical probes for cell biology. After resuspending them in the ASSAY buffer (a SPAchip®-to-cell ratio of 2:1 is recommended), an overnight incubation is required to allow chips to be incorporated in the cytosol. SPAchip® will remain in the cytosol for longer than one month to monitor the evolution of the culture.
Kit box contains:
1 ASSAY SPAchip® eppendorf tube (Quantity: 5 x10^6 ASSAY SPAchip®)
1 Control SPAchip® eppendorf tube (Quantity: 5 x10^5 CONTROL SPAchip®)
1 ASSAY buffer tube (Quantity: 10mL)
Quick start guide
DATA SHEET
APPLICATION NOTE
User protocol
Key features
- SPAchip® assay kits are novel cell-based assays for living single-cell that bring together the fields of nanotechnology and cell biology.
- SpheroCHECK SPAchip® pH Green Single-Detection Kit enables continuous, simultaneous, and accurate monitoring of intracellular and extracellular pH in living 3D models.
- SpheroCHECK SPAchip® monitors pH changes, facilitating an effective quality control of living cells to warrant the standardization and reproducibility of your cell experimental methodologies.
- SPAchip® is compatible with other detection technologies used for numerous spheroid assays; therefore, it can be multiplexed with other stainings for imaging of 3D cultures.
- SpheroCHECK SPAchip® enables a more comprehensive study of spheroid formation and physiology.
- SPAchips remain in the cytosol reluctant to exocytosis during spheroid manipulation. In adittion, SPAchips can be internalized by cells prior to, along or after formation of spheroids.
- SPAchips allows for cell tracking of single cells after spheroid dissociation and single-cell recovery.
- pH fluctuations directly affect cell viability, proliferation rates, and metabolic activities, ultimately affecting spheroid formation and condition. Thus, our technology allows to accurately assess the efficacy and toxicity during drug testing in 3D cell culture.
Technical specifications
| Product code | SS-001-PHG |
| Amount | ~5 millions of SPAchips |
| Applications | Cell Viability, proliferation, cell image acquisition |
| Assay Time | 30 minutes |
| Solubility | Soluble in assay buffer (aqueous) |
| Fluorescence | λex:488 nm; λem: 520 nm |
| Detection method | Green Fluorescence |
| Assay type | Living single-cell based |
| Platform | Fluorescence microscopy, HCS/HCA platforms and flow cytometry |
| Sample type | 3D structures: Spheroids. |
Image Acquisition and Quantitative Analysis
SpheroCHECK SPAchip® kits are designed to dynamically shift fluorescence intensity values in response to intracellular changes in analyte concentration. Fluorescence intensity is measured by exciting the live cell sample at 488 nm and detecting fluorescence emission at 520 nm. Relative intensity units depend on the instrumental setup (excitation power, detector efficiency, etc.). To obtain quantitative data, a calibration curve needs to be analyzed in the same run. Data acquisition can be adapted to fluorescence microscopy and flow cytometry.
Figure 1: a) Emission spectra of SpheroCHECK pH Green probe when excited at the indicated wavelength. b) Brightfield image of SPAchips® once in solution (Image from Torras et al. 2015 DOI: 10.1002/adma.201504164), Scale bar = 10 µm.
SPAchip® fluorescence signal quantification can be adapted to any image analysis software. Object segmentation should be performed to identify SPAchips inside of the cellular region in every sample. Fluorescence intensity units can be exported as a spreadsheet to plot graphical summaries of the results. To quantify intracellular pH, use the fluorescence of the calibration wells to plot fluorescence units vs calibration pH. Interpolate the values of the sample wells to obtain the intracellular pH value of the sample wells.
* Contact A4cell staff for assistance with imaging setup and data analysis. Ask for our cell biology image analysis guide at info@a4cell.com
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