Tunnelled haemodialysis catheters (t-HDC) are prone to colonization by microorganisms, resulting in increased morbidity and mortality. A previous study concluded that all culture-negative catheters removed from cancer patients were colonized by microbial biofilms when examined by scanning electron microscopy (SEM). Examination of t-HDC by SEM has not been published before. A total of 44 segments (0.5 cm each) from 11 ex-vivo t-HDC were examined by SEM prior to endoluminal brushing and quantitative culture to determine their colonization status. Endoluminal brushing yielded a positive culture from two catheters. Methicillin-sensitive Staphylococcus aureus was grown from one catheter and a Streptococcus species was cultured from the second. SEM examination revealed universal endoluminal coverage by adherent biological material (ABM), which was composed of fibrin, platelets and other host-derived products. However, bacterial cells were visible on the two culture-positive catheters and on two out of nine culture-negative catheters, and were possibly present on one culture-negative catheter. In conclusion, in this study the prevalence of microbial colonization of ex vivo t-HDC was 18% using the endoluminal brushing technique and 36% when examined by SEM. The previously reported universal microbial colonization of central venous catheters is likely to represent coverage by ABM rather than by microbial biofilms.