Acoerela Dye Platform

The World's First Full Bilayer Spanning Lipophilic Dyes

Water Soluble Acoerela Dyes

Many dyes are highly hydrophobic and often require reconstitution in an organic solvent such as DMSO or ethanol. Unfortunately, these solvents may induce artifact changes when present in biological samples because they are not as biocompatible as aqueous buffers. Moreover, some dyes (e.g., carbocyanine) have the tendency to form nanoaggregates when they are being serially diluted from their DMSO or ethanolic stocks. These dye nanoaggregates are potentially sources of false positive signals when analyzing small biological particles like exosomes.

Why choose Acoerela

Fluorogenic

Acoerela dyes are characterized by a high quantum yield (>25%). The significant increase in fluorescence is only observed when the dyes are bound to their target, in which there is change in the hydrophobicity of the environment surrounding the dyes. The stark difference in emission profiles between bound and unbound dyes underlie the “light on” mechanism.

Diverse color options

The current chromophore platform of Acoerela dyes allows us to derive up to 3 different colors

and we are expanding on the excitation/emission profiles of these dyes to enable more multiplexing possibilities in different applications.

Water soluble

Acoerela Dyes are highly water soluble and do not form micelles or nanoparticles when reconstituted in aqueous buffer, unlike that of the family of PKH26, DiR and DiD dyes which will give rise to false positives in flow cytometry analysis of exosomes.

Extracellular Vesicles Dyes

The highly water soluble Acoerela Extracellular Vesicles Dyes enable the accurate detection of nanosized extracellular vesicles, exosomes and liposomes, without any artifacts or false positive signals from the dye itself. This is because of the unique patented chemistry of the Acoerela dyes that prevent the dye from self-assembly into micelles or nanoparticles. The Acoerela dyes are soluble in aqueous buffer (e.g., water, PBS, saline) at >2mg/mL concentration. These dyes are highly emissive and only fluoresce when bound to the target. They have been validated with exosomes obtained from MSC, PC3, HT29 and A549 cell lines.

Key Features

Flow cytometry analysis of exosomes labelled with Acoerela Extracellular Vesicles Dye.

Dot plot of (a) unstained exosomes, (b) exosomes labelled with 1 µM Acoerela Exosome Dye, (c) 1 µM Acoerela Exosome Dye only and (e) 1 µM of commercially available PKH26 dye only i.e., in the absence of exosomes.

Imaging Flow Cytometry

PC3 exosomes were labeled with Acoerela Extracellular Vesicles Dye and purified using the Amicon 100KDa filter before they were added to A549 cells, incubated, and analyzed on Imagestream (Amnis) after (a) 2 h, (b) 4 h and (c) 8 h of incubation. Acoerela Extracellular Vesicles Dye-labeled exosomes were observed as spots (green). In the absence of exosomes, no fluorescence was observed in the cells because all the free dye were removed (d). By contrast, micelles of DiD dye were not purified and thus there was still fluorescence observed from the cells, suggesting the uptake of the dye aggregates which can be mistaken for DiD-labeled exosomes instead.

Bacteria Gram-Selective Dyes

Bacteria is usually either identified as Gram-positive or Gram-negative depending on its membrane structure. Gram-negative bacteria are typically characterized by the presence of an outer membrane that comprises of highly charged lipopolysaccharides (LPS), which is absent in the Gram-positive counterparts.

The Acoerela Bacteria Gram-Selective Dye takes opportunity of this difference in the cell wall of the two Gram-types, and can be used to selectively label the Gram-positive bacteria due to the more rapid entrance of the dye into the cell. The selectivity of the Acoerela Dye has been tested in mixed populations of Gram-positive bacteria (Staphylococcus aureus, Enterococcus faecalis) and Gram-negative bacteria (Pseudomonas aeruginosa, Escherichia coli).

In situ Gram-typing of a mixed microbial biofilm using Acoerela Dye and a counter stain. Confocal microscopy images of E. faecalis (Gram-positive, coccus-shaped) and E. coli (Gram-negative, rod-shaped) polymicrobial biofilm after subsequently stained with a gram-positive selective dye and a counter-stain for gram-negative bacteria. (A) Merged images from both channels of (b) Acoerela Gram-Selective Dye channel (Ex405/Em450-490) and (c) FM 4-64 channel (Ex639/Em640-700).

Key Features

Membrane Dyes for Mammalian Cells

Cell division monitoring is an important part of cell biology research on immune regulation, cell function, quiescence, proliferation and differentiation. Fluorescence cell division trackers are often used in flow cytometry and imaging to study the interactions and fates of different cell types in vitro and in vivo. The Acoerela Membrane Dyes can be used to track cell division due to their stable binding and low cytotoxicity towards mammalian cells. These dyes also have very high quantum efficiency, that allow fluorescence tracking over long periods of time.

High quantum yield (>25%)
Stable binding to mammalian membrane
Low photobleaching
Low cytotoxicity
Suitable for cell division tracking and labelling red blood cells
Variety of colors available

Acoerela cell membrane dye

Confocal images of HepG2 cells that were co-stained with Acoerela Membrane Labelling Dye and a commercially available membrane dye, FM 4-64. (a) Acoerela dye channel (Ex405), (b) FM 4-64 channel (Ex488), (c) brightfield channel, (d) merged channel. The co-localization of Acoerela Membrane Dye and FM4-64 demonstrates the membrane specificity of the Acoerela Dye.

Acoerela cell division tracking

A549 mammalian tumor cells labeled by Acoerela Cell Division Tracking dye. Acoerela Dye-labeled cells (red) have a 3-log fold increase in fluorescence intensity compared to the unstained cells (black) were maintained over 4 passages through serial passaging before analysis on CytoFLEX (Beckman Coulter). Orange: 1st passage, Green: 2nd passage, Blue: 3rd passage. Magenta: 4th passage.

Red Blood Cell Labelling

Flow cytometry analysis of samples that have a mixture of unstained red blood cells (RBC) and RBC stained with Acoerela dye, in the ratio of (a) 0:5, (b) 2:3, (c) 3:2 and (d) 5:0. The Acoerela dye for RBC remains stable in the cell after labelling and does not cross-over into samples that are not intended to be labelled with the dye.

NIR-II Acoerela Dye

Near-infrared II (NIR-II) dyes have been gaining traction in the recent years due to the ability of the light to better penetrate deeply into tissues for imaging, and the low autofluorescence and background signals at that region of the electromagnetic spectrum. The Acoerela NIR-II dye allows the tracking of tumor growth inside a mouse model for an extended period of time, without losing significant emission. The long-lasting fluorescence capabilities and relatively high quantum yield enables researchers to study tumor progression and also the effect of drug treatment regimes against cancer in vivo, which can potentially reduce the number of tests mice, that are traditionally sacrificed at regular time intervals to characterize the tumor.

Acoerela cell membrane dye

NIR-II Acoerela Dye. The NIR-II Acoerela Dye labelled (a) mammalian cells are differentiated from the unstained control cells by flow cytometry (CytoFLEX, Beckman Coulter)

In vivo imaging.

(Top) In vivo fluorescent images of the intracranial tumor over 14 days and the subcutaneous tumor over 26 days (bottom) by collecting emission from the NIR-II Acoerela Dye. In both tumors, the fluorescence intensity of the NIR-II Acoerela increased as the tumor grew over time, allowing for a non-invasive method for tracking tumor growth without repeated sacrificial of the mice.