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PromoKine Cell Biology catalog!
Scientifically profound & practically
oriented off-the-job training
Benefits
Applications
Fluorescent dyes offer extraordinary sensitivity and extended possibilities for the visualization and analysis of intracellular processes and cell structures such as the cell membrane, mitochondria, the nucleus, and the cytoskeleton. Some dyes are also suited for staining only viable or dead cells. That is an important feature for elucidating the effects of certain agents on cell viability.
PromoKine provides a select choice of reagents for staining different cell organelles and structures as well as for determining processes like membrane potential changes. Some of these reagents are available as esters, which pass through the membranes of viable cells. Once inside, the ester group is hydrolyzed by cellular esterases, and the dye can accumulate in the cell. Some dyes are non-fluorescent in their ester form but fluoresce strongly when the ester group has been removed.
Applications
Benefits
The Live/Dead Cell Staining Kit I utilizes two fluorescent dyes, Calcein-AM and Propidium Iodide. Calcein-AM penetrates the cell membrane of living cells. Once inside, the cellular esterases present in living cells hydrolyze it to cell membrane-impermeable, green-fluorescent Calcein (emission maximum at 515 nm). In contrast, Propidium Iodide (PI) can only pass through the disrupted membrane areas of dead cells where it intercalates with the DNA of the nucleus, emitting red light at 617 nm. Since both Calcein and Propidium Iodide can be excited at 490 nm, a simultaneous monitoring of viable and dead cells is possible using a fluorescence microscope. At an excitation of 545 nm, the number of dead cells can be monitored.
The Live/Dead Cell Staining Kit II also utilizes two fluorescent dyes, Calcein-AM and Ethidium homodimer III (EthD-III). As Propidium Iodide, EthD-III can only pass through the disrupted membrane areas of dead cells where it intercalates with the DNA of the nucleus, emitting red light at 620 nm. Compared with PI, EthD-III is more stable and emits brighter fluorescent signals. Since both Calcein and EthD-III can be excited at 490 nm, a simultaneous monitoring of viable and dead cells is possible as well using a fluorescence microscope. At an excitation of 545 nm, the number of dead cells can be monitored.
The Bacteria Live/Dead Staining Kit enables a two-color fluorescent staining on both live bacteria (green) and dead bacteria (red) using the two nucleic acid dyes DMAO and EtD-III. DMAO is a green-fluorescent dye which stains both live and dead bacteria with intact and damaged cell membranes while the red-fluorescing EtD-III only stains dead bacteria with damaged cell membranes. The kit is applicable to most bacteria types and suitable for use with fluorescence microscopes and flow cytometers.
PromoKine's Fluorescent Cytoplasmic Membrane Staining Kits are ready-to-use dye delivery solutions that can be added directly to normal culture media to uniformly label suspended or adherent culture cells. In addition, NeuroDiO, an improved version of DiO, further improves cytoplasmic membrane labeling by a green fluorescent carbocyanine dye. PromoKine also offers DiB, the first blue cytoplasmic membrane labeling dye. PromoKine's Fluorescent Cytoplasmic Membrane Staining Kits include cytoplasmic membrane orange labeling (DiI), cytoplasmic membrane green labeling (NeuroDiO), cytoplasmic membrane red labeling (DiD), and cytoplasmic membrane blue labeling (DiB). They allow cell populations to be marked in distinctive fluorescent colors for identification after mixing. Double labeling can identify cells that have fused or formed stable clusters.
PromoKine's Nerve Terminal Probes are a series of fluorescent cationic styryl dyes developed to follow synaptic activities at neuromuscular junctions or synapses. These dyes typically have a lipophilic tail (two carbon chains) at one end and a highly hydrophilic, cationically charged head group at the other end.
These nerve terminal probes were originally called FM dyes, and now they are available from PromoKine under own trademark names depending on the wavelength of the dyes (SynapseGreen and SynapseRed).
We also offer a series of fixable nerve terminal dyes, known as AM dyes, which are closely related to the FM dyes. The primary difference between the fixable and nonfixable dyes is that the formers have an additional amino group attached to the positively-charged head group of the dyes. The amino group reacts with formaldehyde fixative, thus crosslinking the dye with membrane proteins. AM2-10, AM1-43 and AM1-44 are derivatives of SynapseGreen dyes that have relatively shorter wavelengths, while AM4-64 and AM4-65 are based on SynapseRed dyes that have relatively longer wavelengths.
When using these nerve terminal dyes, one frequent problem is the background fluorescence due to nonspecific membrane staining. Although most of the background fluorescence can be removed by repeated washings with a buffer, the problem is still significant with dyes that have a longer tail or more double bonds, particularly when the dyes are used in tissue preparations. To reduce the background fluorescence, we offer three quencher or dye-clearing agents: ADVASEP-7, a sulfonated cyclodextrin, forms a water soluble inclusion complex with SynapseGreen C4, thus leaving the dye in the aqueous phase. The dye-dextrin inclusion complex is actually much more fluorescent than the free dye in solution. However, repeated washings remove the inclusion complex and thus lower the background fluorescence. Our unique quencher, SCAS, reduces background fluorescence as soon as it is added to the preparation without the need for repeated washings. Sulforhodamine 101 has also been used to reduce SynapseGreen C4 background staining via fluorescent resonance energy transfer (FRET).