Co-localization

Microscopy is used to measure trafficking of molecules to sub-cellular compartments or the co-localization of proteins on, within, or between cells. The ImageStream system provides an improved method of studying co-localization of probes by combining the automatic and rapid collection of images with the objective quantification of probe location and intensity. The correlation analysis built in to the IDEAS image analysis software provides robust statistically significant results on large numbers of cells so that phenotypically distinct subsets of cells can be analyzed.

Co-localization of CpGB to lysosomes and endosomes in primary plasmacytoid dendritic cells

Binding, internalization, and colocalization to lysosomes of CpGB by plasmacytoid dendritic cells (pDC) is measured using the ImageStream. In combination with immunophenotyping, spatial resolution and intensities of lysosomes (green) are correlated with CpGB (red) in pDC (orange). This data highlights several strengths of the ImageStream: 1) combination of immunophenotyping (pDC identification via fluorescent intensity staining) with image correlation and internalization analysis, allowing measurement of colocalization and internalization of molecules within specific subsets. 2) ability to measure these events in small, primary cells with small cytoplasmic compartments. For the compete time course of endosome and lysosome co-localization, see the application note.

Co-localization of an antibody-drug conjugate to endosomes or lysosomes

In this experiment we use the capabilities of the ImageStream system to measure the transit of a fluorescently labeled antibody-drug conjugate (ADC) through the cellular endocytic pathway. The high spatial resolution afforded by the ImageStream system allowed accurate measurement of ADC co-localization to endosomes and lysosomes. See the application note for more details.