Haufen, i.e. discrete three-dimensional cast-like polyomavirus aggregates in the urine, mark polyoma BK-virus nephropathy (BKN) with positive and negative predictive values of greater than 95%. They are novel diagnostic biomarkers of BKN, an important infectious complication post kidney transplantation. Here, we describe technical details of Haufen detection by negative staining electron microscopy. We studied more than 400 urine samples from over 180 patients and developed easy-to-follow protocols for optimal specimen preservation and preparation, including sample clarification and concentration. We detail diagnostic clues to detect Haufen and illustrate pitfalls, including "Haufen-look-alikes," which can hamper the interpretation. Urinary Haufen detection by negative staining electron microscopy is a new application of an old, well-established diagnostic technique. The protocols described here are useful for pathologists and electron microscopists to search for Haufen in voided urine samples, to predict BKN noninvasively, and to help clinicians managing renal allograft recipients.