In vivo localisation assays. Subcellular localisation of peroxisomal marker enzymes demonstrate that mutations within the ancillary SCP2 binding site of Pex5p do not impair peroxisomal import. Pex5p-deficient fibroblast cells were co-transfected with a plasmid expressing GFP-SCP2 either with (A) or without its PTS1 sequence (B) and plasmids expressing wild-type (WT) or mutant forms of Pex5p, carrying the point mutations R608W, D624W or S600W. Forty eight hours after transfection GFP-SCP2 was visualised by direct fluorescence (green colour), while Pex5p (A) and Pex14p (B) were detected by immunofluorescence microscopy. Scale bars (in bottom right micrographs) indicate 10 μM.