technology comparison: flow cytometry, ELISA, FMAT

flow cytometry versus mirrorball

Flow cytometry analysis requires an additional step to suspend adherent cells. This often causes shear stress which may result in cells clumping and cause clogging, requiring user intervention to resolve and causing system downtime. TTP Labtech’s mirrorball allows you to screen both adherent and suspension cells in situ thus avoiding such issues.
Traditional cell labelling involves multiple wash steps which are time-consuming and wasteful of reagents. mirrorball is mix and read enabled: so you just add the reagents, read the plate and get your results.
In automated flow, samples are combined into a continuous stream separated by air bubbles. Broken-up air gaps can have serious implications for sample identification and integrity. Sample integrity is at the heart of analytical efforts. With mirrorball you are guaranteed it, as each sample is analysed in situ.
In automated flow, once a sample has been read it is deployed to wastte, so timecourse assays are not possible. mirrorball enables timecourse assays, as non-destructive scanning allows samples to be re-read
Flow cytometry provides data in scatter charts but no visualization of cells. mirrorball generates TIFF images, if wanted. These provide an additional way to visualize the assay, boosting confidence in the data obtained.
Flow cytometry analysis requires a high number of cells (typically 200,000 per test) microplate cytometry uses far fewer cells (down to 2,000 cells per well), which is a large cost saving. It also optimises the use of expensive, biologically-relevant primary cells.

ELISA versus mirrorball

ELISA mirrorball
ELISA usually requires soluble protein fragments or purified antigen, rather than cells. These are less biologically relevant than whole cells. TTP Labtech’s mirrorball can screen intact cells, allowing you to look at antigens in their natural conformation and preserve target epitopes. Using native antigens early in the screening process reduces the likelihood of false positives and may identify true positive hits that may be incorrectly rejected by ELISA.
To produce soluble protein fragments or purified antigen is labour intensive and requires complex automation. mirrorball uses cells expressing the target, so you don’t have to do the work of making purified protein.
The ELISA process requires around 15 wash steps. ELISA conversion to a bead-based format on mirrorball is straightforward. These can be run as a ‘no wash’ assay, which offers significant process efficiencies.
Multiple wash steps in the ELISA process mean low affinity biologics are likely to be missed. Screening in mirrorball’s no wash format is more likely to identify low affinity biologics.
ELISA assesses one target attached to a plate so multiplexing is not possible mirrorball is a 3 laser system with 5 data collection channels, offering multiplexing. This ability to combine multiple assays into one gives decision-making data faster, and at lower cost.


FMAT versus mirrorball

FMAT mirrorball
For the ABI 8200 (FMAT) to identify antigen-specific binding against parental and transfected cell lines requires a separate assay to be run for each cell line, so two wells are needed for each sample. mirrorball’s multiplexing capabilities allows these tests to be performed in a single well, which has the advantages of eliminating inter-plate variability and increasing throughput by halving the number of plates needed.
If antibody binding is low, then object recognition is compromised using the ABI 8200. Even when there is low antibody binding, as adding a counterstain allows mirrorball to reliably detect cell numbers irrespective of antibody staining.
While the ABI 8200 system has been a workhorse in many labs, the instrument is no longer made and support has been discontinued. mirrorball is a proven product, available since 2010. TTP Labtech has an enviable reputation for service and support going back over the last 15 years.
The ABI 8200 is a single laser system that tends to be limited to detecting one colour only mirrorball is a 3 laser system with 5 data collection channels, offering greater multiplexing possibilities. This ability to combine multiple assays into one gives decision-making data faster, and at lower cost.
Scatter detection of beads is not possible using the ABI 8200. mirrorball’s scatter detection of beads offers more sensitive and robust data, as beads can be detected even if no binding takes place.