Analysis of proteinuria: reference limits for urine excretion of albumin, protein HC, immunoglobulin G, k- and Δ-immunoreactivity, orosomucoid and α₁-antitrypsin

Abstract

Efficient use of assessment of urine protein excretion in nephrological practice requires adequate reference intervals. To determine the upper reference limits of urine albumin, protein HC (alpha 1-microglobulin, immunoglobulin G (IgG), orosomucoid (alpha 1-acid glycoprotein, alpha 1-antitrypsin, and kappa- and lambda-chain immunoreactivities, the concentrations of these proteins were measured in urine samples from 95 healthy, adult individuals, using rapid, generally available methods and with conditions for urine collection which secured stable protein levels. The obtained values were expressed in mg 1(-1), as the urine protein-creatinine index and as fractional protein-creatinine clearance. No differences were found between the upper reference limits in the first voided morning urine samples and the randomly collected urine samples, nor between the upper reference limits in urine samples collected from males and females. The urinary excretion of the tested proteins did not correlate to age, positive dipsticks for haematuria nor to granular casts in urine sediment. Thus, the same upper reference limits can be used for both sexes and regardless of the type of urine collection. The upper reference limits of urine protein-creatinine index found in this study were: for albumin, 3.8 mg mmol(-1); for protein HC, 0.7 mg mmol; for IgG, 0.8 mg mmol(-1); for orosomucoid, 0.7 mg mmol(-1); for alpha 1-antitrypsin, 0.2 mg mmol(-1), and for kappa-immunoreactivity 0.7 mg mmol(-1). The upper reference limit for lambda-immunoreactivity was below the detection limit.

Reference intervals for 24 laboratory parameters determined in 24-hour urine collections

“…These concentrations were replaced by LLOQ divided by the root square of two [12] for statistical analysis as well as the calculation of reference intervals. The reference interval obtained for IgG using this approach is very close to that of the aforementioned study [21] with a similarly large 90% confidence interval for the upper reference interval limit. The concentrations of transferrin and of α2-macroglobulin were above the corresponding LLOQ in only three and only in one out of 241 samples, respectively, i.e.…”

“…albuminuria [19], have been defined by consensus of experts on the basis of clinical outcomes in epidemiological or clinical intervention studies rather than on the frequency distribution in the population. Similarly to another reference interval study for proteinuria marker proteins [21] the concentrations of these proteins were below the LLOQ of the analytical methods for many of the urine samples of our study. These concentrations were replaced by LLOQ divided by the root square of two [12] for statistical analysis as well as the calculation of reference intervals.…”

Coelution of Other Proteins with Albumin during Size-Exclusion HPLC: Implications for Analysis of Urinary Albumin

“…The aggregate concentration in plasma of globulins with a molecular size similar to albumin, proteins such as transferrin, ␣ 1 -acid glycoprotein, ␣ 1 -proteinase inhibitor, ␣ 1 -antichymotrypsin, ␣ 2 -HS glycoprotein, hemopexin, Gc-globulin, and many others (31 ), in healthy individuals is ϳ25% that of albumin; this aggregate concentration is even higher during acutephase reactions (32 ). Because the passage of plasma proteins through the glomerular barrier into urine is primarily a size-dependent process with a size-exclusion limit approximately the size of albumin (33 ), it is not surprising that all of the albumin-sized plasma components are also found in urine (15)(16)(17)(18)(19)(20)(21)(22)(23)(24)(25)(26)(27)(28)(29)(33)(34)(35)(36)(37)(38)(39). There is considerable variation in the reported proportions of individual globulins relative to albumin in urine (Table 3).…”

“…However, the aggregate concentration of albumin-sized globulins in urine appears to be ϳ20% that of albumin, with significant variation in different physiologic states. (21 ) a Proteins are grouped as those that coelute with albumin during size-exclusion HPLC and proteins that potentially overlap with albumin during this analysis. b Ratio of 95% limit of protein vs 95% limit of albumin.…”

“…Although albumin is the most abundant plasma protein, there are many other plasma proteins of approximately the same molecular size, such as transferrin, ␣ 1 -proteinase inhibitor, hemopexin, ␣ 1 -acid glycoprotein, ␣ 1 -antichymotrypsin, Gc-globulin, ␣ 2 -HS glycoprotein, and transthyretin, that are major urinary components (15)(16)(17)(18)(19)(20)(21)(22)(23)(24). Fundamental principles suggest that size-exclusion HPLC will not resolve albumin from other urinary proteins of similar molecular size.…”