We have compared two in vitro methods and three variations of kidney background (BG) subtraction within a gamma camera method (41 examinations, 31 patients) for determination of effective renal plasma flow (ERPF) using 131I orthoiodohippurate (OIH). Method I: plasma samples at 20 and 45 min after OIH injection, ERPF = dose X slope/intercept; Method II: 45-min plasma sample, ERPF = -51.1 + 8.21x + 0.019x2, x = dose/45-min plasma activity/I. Individual kidney and total ERPF were determined from gamma camera (GC) methods using renal uptake 1-2 min after injection. All methods were compared against Method I (previously validated against paraaminohippurate (PAH) clearances). Method II, which requires one blood sample is more accurate than GC methods. GC methods are insensitive to operator variability in placement of renal and BG regions of interest. They may be useful to follow changes in relative or total ERPF, but accurate depth correction of renal data is suggested. In vitro, blood sample-based methods are more accurate.