EMBL

last update

09 Feb, 2005

Assays

The grouping of large sets of proteins according to subcellular localisation allows functional assays to be designed that are tailored to this information. For example we are particularly interested in membrane traffic, and therefore can use the localisation information to specifically concentrate on  the subset of novel proteins that we find localising to relevant membranes such as the endoplasmic reticulum and Golgi complex. In this way we can more systematically screen the novel proteins for the function of interest - for example secretion.

This has been facilitated by the development of automated microscopy platforms that can acquire large data sets for quantitative image analysis. Quantification of fluorescence from multiple channels can be made on a cell-by-cell basis, and a wide variety of parameters can also be measured. Below is listed some of the features that such an analysis can provide us, and the figure shows an example image acquired on an automated microscope, and its subsequent quantification.

1) Active autofocus / quality control (cell orientated)
2) Pattern based mask generator
3) Pattern based auto-threshold
4) Pattern / feature based cell separator
5) Cell statistics (number, size, shape etc.)
6) Morphology parameter
7) Background subtraction

Microscopy-based functional assays and automated data analysis