The LeviCell contains 2 LED channels which can be used to visualise flourophores in your samples
When performing Live Cell Enrichment on the LeviCell, you have the flexibility to switch the light source to one of the built-in LED sources. This feature allows you to easily visualize any compatible flourophores, stains or dyes that are present. The LEDs available to you have specific excitation and emission spectra. The Green Channel LED has an excitation wavelength of 474 nm and an emission wavelength of 524 nm, while the Red Channel LED has an excitation wavelength of 560 nm and an emission wavelength of 628 nm. By using the LEDs, you can easily detect and analyze the different fluorescent signals emitted by the samples, which can provide valuable insights into the biological processes happening within the cells.
Note: Download and read the full application note here.
Table 1. The LED light sources and their respective excitation / emission wavelength.
| LED source | Excitation (nm) | Emission (nm) |
| Green Channel | 474 | 524 |
| Red Channel | 560 | 628 |
What dyes can I visualize using these channels?
The LeviCell platform has undergone extensive testing with a variety of dyes and probes, and the results have been compiled in Table 2 for your reference. While the table is not exhaustive, it provides a comprehensive list of the dyes and probes that have been used and visualized in the LeviCell.
Table 2. Dyes, probes and fluorescent proteins with optimal concentrations as tested on the LeviCell platform. Ex=Excitation, Em=Emission, WS=Working Solution, [WS]=Working Solution Concentration. Levitation buffer is composed of the media or buffer plus the levitation agent. Levitation Buffer volume = 300 µL.
| Dyes/Probes/ Flourescent Proteins |
Channel Ex /Em |
Protocol | Use Cases | Mechanism |
| Propidium Iodide |
Red Ex 483/Em 636 |
WS: 1 mg/mL Use 1:300 into the Levitation Buffer + cells |
Detection of dead cells and nuclei | DNA intercalating dye which is permeable to cells with compromised membrane only |
| Acridine Orange (AO) |
Green Ex 500/ Em 526 |
WS: 0.2 mg/mL Use 1:100 into the Levitation Buffer + cells | Detection of all nucleated cells | Cell nucleic acid binding dye that emits green when bound to dsDNA |
| Calcein AM |
Green Ex 495/ Em 515 |
WS: 5 µM Resuspend 1 million cells into 100 µL of WS. Incubate 15 min, spin and resuspend with the levitation buffer. | Detection of viable cells | Passive diffusion of dye. Inside live cells calcein AM is hydrolysed to green-fluorescent calcein by intracellular esterases. |
| Fluorescein Diacetate (FDA) |
Green Ex 498/Em 518 |
Follow vendor’s staining recommendations | Visualization of live protoplasts and pollen | Passive diffusion of dye. Inside live cells it is deacetylated to green-fluorescent fluorescein by intracellular esterases. |
| FITC Annexin V |
Green Ex 488/Em 517 |
Follow vendor’s staining recommendations | Detection of apoptotic cells | During early apoptosis Annexin V is exposed to the extracellular membrane and the probe recognizes it. |
| FITC-conjugated antibody, eg. anti-CD45 FITC |
Green Ex 488/Em 517 |
Incubate with anti-CD45 FITC (1:10 dil) for 30 min at RT | Detection of cell surface protein CD45 | CD45 is a glyoprotein expressed on the cell surface of all nucleated hematopoietic cells |
| MitotrackerTM green FM |
Green Ex 490/ Em 516 |
WS: 200 nM Use phenol red free media to incubate cells during 45 min with the dye, spin the cells and resuspend with levitation buffer. |
Detection of live cells | Passive diffusion of dye. It accumulates in active mitochondria in a potential-dependent manner. |
| Cell TrackerTM Red CMTPX |
Red Ex 577/Em 602 |
WS: 10 mM Resuspend 1 million cells into 200 µL of WS. Incubate 15 min at 37 C, spin and resuspend with levitation buffer |
Detection of stained cells in a mixture | Passive diffusion of dye. It is retained in living cells through several generations. |
| FluoroMyelinTM Green |
Green Ex 479/Em 598 |
Follow vendor’s staining recommendations | Visualization of myelin | It binds to myelin protein of brain digestions |
| mCherry |
Red Em 585/Ex 611 |
Detection of cells that have been successfully transfected using fluorescent protein expression marker | Expression in a cell | |
| Green Fluorescent Protein (GFP) |
Green Em 498/Ex 509 |
Detection of cells that have been successfully transfected using fluorescent protein expression marker | Expression in a cell |