The temporal model of IgG subclass switching supports sequential IgG conversion; however, the diversity of previously stimulated B cells is known to shape responses in different directions. Recent studies suggest that different subclasses of IgG (1-4 in humans) possess varying capacity to cause graft injury. As IgG subclass switching is a dynamic process, we performed a preliminary exploratory study in 23 adult and pediatric recipients (14 kidney/3 kidney-pancreas/3 heart/2 kidney-liver/1 pancreas) to investigate the kinetics and relative magnitude of donor-specific antibodies (DSA) IgG subclasses and compare to total IgG over time. Serial post-transplant samples were tested for total IgG, IgG3 and IgG4 subclasses using Class I and II Luminex SAB. All patients had total IgG DSA >1000 MFI at all time points tested and most were receiving treatment for active AMR.

Total IgG antibodies (Ab) were compared with those detected by IgG3 or IgG4 reagents for each HLA Ab specificity. Interestingly, in 12 patients, some Ab directed at the same target were positive for both IgG3 and IgG4 in a single serum sample.

In 21 patients, MFI values for the positive total IgG beads were higher or comparable to those seen with IgG3 or IgG4 reagents. However, 2 patients showed much higher MFI values for the IgG4 reagents than the total IgG (HLA-C Ab in 1 patient and HLA-DQ Ab in the other). While most patients had noticeable IgG3 or IgG4 subclass binding in at least one serum sample, the MFI varied dramatically over time, and generally followed a similar pattern of increase or decrease as the corresponding specificity in the total IgG assay. Notably, we did not see class switching over time from IgG3 to IgG4 for individual Ab, although this was not a comprehensive study. Among the 10 patients that were tested at the time of active AMR with C4d+ biopsies, only 2 had IgG3+ DSA and 3 had IgG4+ DSA.

In summary, MFI of IgG subclasses fluctuate over time and do not seem to correspond to the clinical status of the patient. Some Ab may be positive for both IgG3 and IgG4, which can complicate analysis in clinical studies. We suspect that the reagents currently available are not ready for clinical use, and we advise significant caution when interpreting results, especially those obtained at a single time point.

CITATION INFORMATION: Pinelli D., Gopalakrishnan M., Joong A., Thrush P., Tambur A. Temporal Variability of IgG Subclasses Am J Transplant. 2017;17 (suppl 3).

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