Measurement of Serum Levels of Complement Component 4d as a Quantitative Test of Antibody-Mediated Rejection

Abstract

Complement component 4d (C4d) is a degradation product of the classical pathway. C4d immunohistochemistry is used for the diagnosis of complement-dependent antibody-mediated rejection (ABMR) in posttransplant renal biopsies. We studied serum C4d levels, measured by a noninvasive and quantitative test, in two posttransplant patients (A and B) with worsening renal function and biopsies demonstrating peritubular capillaritis and glomerulitis. Serum samples from patients A and B were obtained within 24 hours of biopsy results indicative of ABMR. The patients' sera and eight controls were stored at -20°C before testing the samples. A commercially available enzyme-linked immunosorbent assay kit was used to measure serum concentrations of C4d (Uscn Life Science Inc, Hubei, China), following the manufacturer's instructions. C4d immunohistochemical staining was performed on formalin-fixed, paraffin-embedded renal biopsy specimens. In addition, the patients' sera were tested for anti-HLA Class I and Class II donor-specific antibodies (DSA) by solid-phase immunoassay (Luminex, Austin, TX). Patient A, with diffuse peritubular capillary C4d reactivity of greater than 75%, had a serum C4d level of 4.4 mg/L. Patient B, with 0% peritubular capillary C4d reactivity, had a serum C4d level of 1.7 mg/L. Both of the patients' serum C4d results were compared to the control group (mean ± SD), 2.4 ± 0.66 mg/L, range 1.6-3.3 mg/L. Patient A had two strong anti-class I DSA (both to the B-loci), and two weak anti-class II DSA (DQB1 and DQA1). Patient B had one moderate anti-Cw DSA. Serum C4d levels of patients A and B were consistent with biopsy C4d staining and presence of DSA. The measurement of serum C4d levels may serve as a quantitative test of complement-dependent ABMR in renal transplant recipients.