Nephrotic-range proteinuria (NRP), defined as urinary protein excretion ≥3.5g/day, represents a heterogeneous clinical entity with divergent pathophysiological mechanisms and clinical presentations.1 While classically associated with nephrotic syndrome (NS) – characterized by hypoalbuminemia (<3.0g/dL), edema, and hyperlipidemia – a substantial subset of patients maintain normal serum albumin despite massive proteinuria, termed nephrotic-range proteinuria without nephrotic syndrome (NRP without NS).2,3

Recent advances suggest that patients with NRP without NS maintain albumin homeostasis through enhanced tubular reabsorption mechanisms, representing distinct pathophysiological processes from full-blown nephrotic syndrome.4–6

The diagnostic landscape for membranous nephropathy (MN) has been transformed by the discovery of anti-phospholipase A2 receptor (PLA2R) antibodies, present in 70–80% of primary MN cases.7 The 2021 KDIGO guidelines represent a paradigm shift, stating that “kidney biopsy is not required to confirm MN diagnosis in patients with nephrotic syndrome and positive anti-PLA2R antibody test”.8 This recommendation, based on studies demonstrating 97–99% specificity and positive likelihood ratios, permits non-invasive diagnosis in 60–70% of MN cases when specific criteria are met.9,10 Although 80% of patients with primary MN present with NS clinical features, approximately 20% of patients present without nephrotic syndrome.11

We aimed to compare anti-PLA2R antibody diagnostic yield between nephrotic syndrome and nephrotic-range proteinuria without nephrotic syndrome phenotypes, to provide evidence for phenotype-based testing algorithms that optimize diagnostic accuracy.

A total of 156 patients with nephrotic-range proteinuria were included in the analysis. Seventy-three patients (46.8%) presented with NRP without NS, while 83 patients (53.2%) had NS. The overall cohort had a mean age of 54.3±16.5 years with 45% female representation.

Baseline demographic and clinical characteristics were comparable between the groups. Age, sex distribution, and body mass index showed no differences. Hypertension was the most frequent comorbidity in both groups, without a statistically meaningful variation. Diabetes mellitus was more common in the NRP without NS group, but the difference was not statistically significant (41% vs 27.7%, p=0.07). Serum creatinine and eGFR did not differ significantly between groups.

Anti-PLA2R antibody positivity was significantly higher in patients with NS compared to those with NRP without NS (26.5% vs 9.6%, p=0.007).

Kidney biopsies were performed in 92 patients (58.9% of the total cohort). Combined membranous nephropathy diagnosis (via kidney biopsy or serology) was significantly more frequent in patients with NS compared to those with NRP without NS (43.3% vs 23.2%, p=0.008). The data is presented in Table 1.

Among the 53 patients diagnosed with membranous nephropathy 36 presented with NS and 17 presented with NRP without NS. Both groups had similar eGFR and creatinine values. Anti-PLA2R antibody positivity rates were numerically higher in nephrotic syndrome patients, though this difference did not reach statistical significance (61% vs 41%, p=0.17). Anti-PLA2R antibody levels were significantly higher in nephrotic syndrome patients compared to those with NRP without NS (157.64 [341.93] vs 27.38 [20.14] RU/mL, p=0.008). The data is presented in Table 2.

In the multivariate logistic regression model including age, sex, diabetes mellitus, proteinuria, and clinical phenotype, nephrotic syndrome remained an independent predictor of anti-PLA2R antibody positivity (aOR=5.00, 95% CI 1.74–14.34, p=0.003). None of the other covariates – including age, sex, diabetes mellitus, or proteinuria – showed a statistically significant association as shown in Table 3.

Our study provides the first systematic evaluation of anti-PLA2R antibody diagnostic yield stratified by clinical phenotype, demonstrating a 2.8-fold higher positivity rate in nephrotic syndrome compared to nephrotic-range proteinuria without nephrotic syndrome. As supported by our multivariate analysis, patients with nephrotic syndrome were approximately five times more likely to be anti-PLA2R positive (aOR=5.00, 95% CI 1.74–14.34, p=0.003), underscoring the strong independent predictive value of clinical phenotype. This finding has important implications for diagnostic algorithms and resource allocation in nephrology practice.

The marked difference in anti-PLA2R positivity between phenotypes likely reflects distinct pathophysiological mechanisms. NRP without NS appears to maintain albumin homeostasis through preserved tubular reabsorption, whereas PLA2R-mediated MN typically causes more extensive glomerular injury, manifesting as full NS. Nonetheless, the identification of MN in 23.2% of NRP without NS patients suggests that a subset may represent early or compensated stages of the same disease spectrum. Consistent with this interpretation, Guerry et al. demonstrated that anti-PLA2R antibodies can be detected before the clinical onset of NS and are associated with a higher likelihood of subsequent progression.15 Conversely, the low rate of anti-PLA2R positivity in NRP without NS may indicate the presence of alternative target antigens such as thrombospondin type-1 domain-containing 7A (THSD7A) or neural epidermal growth factor-like 1 (NELL-1), or other yet unidentified antigens.16,17

Our findings strongly support the KDIGO 2021 guideline requirement for NS presence in non-invasive MN diagnosis protocols.8 However, anti-PLA2R–positive MN can also be observed in the NRP without NS group. This may reflect early or subclinical MN in a subset of patients; in our series MN accounted for 23.2% of NRP without NS cases. While previous studies emphasized secondary focal segmental glomerulosclerosis and diabetic nephropathy in this population2,3 our observation may challenge the traditional view that NRP without NS primarily represents hyperfiltration-mediated disease. If there are no economic constraints, testing for anti-PLA2R antibodies in this group still carries the potential to establish a biopsy-free diagnosis of MN in a subset of patients during initial presentation or follow-up.

Anti-PLA2R antibody titers are well-established markers of disease activity and prognosis in MN.18–21 In our cohort, MN patients with NS had higher proteinuria, lower albumin, and higher anti-PLA2R titers than those without NS, consistent with prior reports. Nevertheless, the presence of anti-PLA2R positivity – even at lower titers – in NRP without NS suggests early or smoldering MN rather than an inactive state. Such patients require longitudinal follow-up, as some may later develop overt NS.22

From a health economics perspective, our findings support phenotype-based testing strategies. Validation studies have demonstrated that anti-PLA2R testing can reliably identify a substantial proportion of membranous nephropathy cases, reducing the need for invasive biopsy and its associated complication risk (0.9–4%), while enabling earlier treatment.23 However, the cost of testing varies across countries and healthcare systems, which may constitute an economic barrier to widespread use.

Several limitations our work warrant acknowledgment. The retrospective, single-center design restricts generalizability, and these findings require validation across diverse populations and healthcare systems. Regional variations in glomerular disease patterns and healthcare resource availability may affect the applicability of phenotype-based testing algorithms. Another limitation was that anti-PLA2R antibodies were measured using the ELISA method. Given that ELISA has lower sensitivity compared to the immunofluorescence assay (IFA) method, our diagnostic sensitivity may have been reduced.18 In addition, the absence of longitudinal antibody kinetics precluded prognostic assessment. Furthermore, some patients biopsy samples lacked anti-PLA2R staining. Finally, no evaluation of alternative antigens (THSD7A, NELL-1) could be conducted. Although known false-positive anti-PLA2R cases were excluded, not all patients underwent kidney biopsy; therefore, residual false positivity cannot be completely ruled out. This may have slightly affected both diagnostic accuracy and cost estimations.

Based on our findings, we propose a phenotype-oriented diagnostic framework. In NS, routine anti-PLA2R ELISA testing remains both clinically and economically justified, as we recently demonstrated.24 In contrast, for patients with NRP without NS, the clinical yield and cost-effectiveness of routine ELISA screening remain uncertain. The complementary use of IFA in seronegative or borderline cases may reduce unnecessary biopsies, but this approach requires prospective health-economic validation. Although we are not aware of literature specifically addressing which NRP without NS patients have MN, MN may be clinically suspected in cases with sudden or progressive proteinuria despite renin angiotensin aldosterone system (RAAS) inhibition, absence of diabetes, and mild hypoalbuminemia. The complete proposed diagnostic algorithm incorporating these criteria is illustrated in Fig. 2.

Fig. 2.

Proposed algorithm for anti-PLA2R testing in NRP. Negative (<14RU/mL), borderline (14–19RU/mL), and positive (≥20RU/mL). ELISA: enzyme-linked immunosorbent assay, IFA: immunofluorescence assay, MN: membranous nephropathy, NS: nephrotic syndrome, NRP: nephrotic range proteinuria, NRP without NS: nephrotic range proteinuria without nephrotic syndrome, PLA2R: phospholipase A2 receptor, RAAS: renin angiotensin aldosterone system.

Clinical phenotype independently predicts anti-PLA2R antibody yield in NRP. When anti-PLA2R testing is economically accessible, broad screening of all patients with NRP may be justified; however, in resource-limited settings, restricting testing to those with full NS provides the best balance between diagnostic efficiency and cost-effectiveness.

Asil Demirezen: 0009-0004-4449-8266

This research did not receive any specific grant from funding agencies in the public, commercial, or not-for-profit sectors.

The authors declare no conflicts of interest.