All the images are of cells labelled with antibodies with fluorescent tags and viewed using a confocal microscope. Confocal microscopes use a laser to scan very thin sections of a cell or tissue sample, giving a much sharper image than is possible on a conventional microscope. The laser scans a square region of the sample and in doing so stimulates any fluorescent molecules (fluorophores) attached to antibodies bound to the sample. The light emitted by the fluorophore is detected with a very sensitive CCT camera and digitised, allowing the image to be saved to disc. Different fluorophores emit different coloured light which can be detected separately by the microscope by using the appropriate filter.
Most of the images presented here are double-label images, meaning two different antibodies with two different fluorophores (green and red) were used on the same sample. Two images are thus obtained: a green image from one antibody, and a red image from the other. If protein X and protein Y are bound to each other in the cell then antibodies to protein X and antibodies to protein Y will label the cell in a near identical pattern, producing a green image that is nearly identical to the red image. If the two images are merged, yellow will be seen where the green and red exactly superimpose. If one image is brighter at a particular point than the other image the colour may not be yellow, but orange or yellow/green. Where there is no colocalisation of proteins/antibodies the combined image will be red or green as in the separate images.
The combining of green and red to make yellow is illustrated below:
