Rapid and sensitive detection of enhanced green fluorescent protein expression in paraffin sections by confocal laser scanning microscopy

Various forms of green fluorescent protein (GFP) have become important repo rters of gene transfer and expression after transfection or infection of ce lls in cell culture. Frequently, molecular biological assays (Northern blot s, PCR) are applied to detect reporter gene expression in target organs. Ho wever, these methods are not suitable for evaluation of tissue- or cell-spe cific expression which would be of great interest especially in case of usi ng tissue-specific promoters. Therefore, organs of transgenic mice with the enhanced green fluorescent protein (EGFP) gene under control of the cytome galovirus (CMV) promoter were processed for histology by formaldehyde fixat ion and embedding in paraffin. Sections were deparaffinized, mounted and ev aluated for fluorescence in a confocal laser scanning microscope. This meth od combines the advantages of direct exploitation of tissue sections withou t further staining procedures with evaluable tissue-, cell-, and even subce llular-specific distribution patterns of EGFP expression in tissues. Result s obtained by direct evaluation of EGFP fluorescence in paraffin sections w ere confirmed by immunohistochemical staining with anti-EGFP. In the presen t report, we demonstrate that application of confocal microscopy on routine ly processed histological preparations is very suitable for determining gen e transfer efficiency and promotor activities.