Co-registration of total internal reflection fluorescence and confocal microscopy images for studying vesicle trafficking in living cells

The dynamics of vesicle trafficking via the cell membrane and the tracking of their subsequent movements within the cell are of great interest in the biomedical sciences. One challenge of image analysis is following the vesicle´s fate continuously from its formation at the membrane to its final destination, as it requires different microscopy techniques to image the complete journey. Total internal reflection fluorescence (TIRF) microscopy is used for imaging events at the cell membrane and laser scanning confocal microscopy (LSCM) is used for imaging the interior of the cell. We present a simple and robust method for co-registration of data sets from the two microscopy techniques. This method is validated on images generated by computer simulation of the image formation process in TIRF and in LSCM. The registration parameters are recovered with error less than 1% in presence of Gaussian noise up to SNR of 3.8dB. Registration of real microscopy data is shown and the accuracy of the retrieved parameters is compared and agrees well with values obtained manually and a difference in squares measure.