PSL Alliance member Mabtech takes us through a study to assess their IRIS FluoroSpot reader
Since our beginning more than 30 years ago, we have been discussing ELISpot, FluoroSpot and immune responses with scientists. We realized that automated readers required calibrations and individual user settings, which often led to subjective interpretation of the spot images.
There had to be a better way – a more scientific way.
Combining our understanding of biological applications and the expertise of Joakim Jaldén of the KTH Royal Institute of Technology in signal processing, we developed a novel spot counting algorithm called RAWspot™ technology which makes spot data collection totally unbiased.
Our aim of the present study was to compare Mabtech IRIS with another commercial FluoroSpot reader requiring user-defined settings, to evaluate whether user-defined input and experience in FluoroSpot analysis influences spot counts.
The Mabtech IRIS FluoroSpot reader has fixed camera settings and RAWspot technology, whereas other commercial readers require user-defined settings of the camera, algorithm and XY-table. To assess the effect of user-defined input on analysis, an external organization was contracted and five operators of varying experience levels were given a FluoroSpot plate with wells in four replicates of an unknown sample. The operators were instructed to analyse the plate using two different readers: (i) Mabtech IRIS User’s expertise level and (ii) a competitor FluoroSpot reader.
Five operators counted an unknown sample in four replicates. With Mabtech IRIS the operators’ average spot-forming units (SFU) ranged between 191-202 spots, resulting in a coefficient of variance (CV) of 3%. With the competitor reader, the operator range was between 24-160 spots (CV 59%). A summary of the results is shown in the graph to the left.
Mabtech IRIS minimizes the user-defined subjective input of the analysis, thereby reducing the influence of the operator on the end result.
Plug-and-play functionality minimizes user-bias
Five operators were instructed to read the same plate on the Mabtech IRIS and a competitor reader.
Accurate spot center detection is key
The RAWspot technology and default settings utilized by Mabtech IRIS accurately detects each spot center and gives precise spot counts.