Table 2.
Summary of different methods for detecting IgG subclasses against HLA and non-HLA targets.
| Reference | Organ/target | Determinants of positive cutoff | Antibody source |
|---|---|---|---|
| Lefaucheur et al. (130) | Renal/HLA | For each IgG subclass antibody and each single antigen bead, the mean MFI values from 4 non-sensitized healthy male controls +5 SD, and if that value was <500 MFI; the IgG 1–4 reactivity was considered positive if the normalized MFI was ≥500 | Southern Biotech |
| Khovanova et al. (129) | Renal/HLA | >5× the greater than the average negative control bead MFI in the single antigen bead assay of all tests for each subclass. IgG1: 120.6, IgG2: 72.0, IgG3: 62.7, IgG4:17.2 | Southern Biotech |
| Lowe et al. (128) | Renal/HLA | >5× the greater than the average negative control bead MFI in the single antigen bead assay of all tests for each subclass. IgG1: 120.6, IgG2: 72.0, IgG3: 62.7, IgG4:17.2 | Southern Biotech |
| Kaneku et al. (127) | Liver/HLA | Normalized trimmed MFI higher than 500 was defined as positive on the basis of binding patterns after validation and dilution experiments | Southern Biotech |
| Honger (126) | Renal/HLA | A positive result was defined by a MFI value above a cutoff that was generated for each IgG subclass and for every individual bead by using four negative control sera from healthy non-sensitized and HLA antibody-negative men: cutoff MFI = mean NC1–4 + 3 SDs NC1–4. To determine the amount of IgG subclasses, we used the ratio above the corresponding cutoff (i.e., ratio = MFI IgG subclass divided by MFI cutoff) | Southern Biotech |
| Jackson et al. (131) | Endothelial cells | Median fluorescence values for IgG subclasses identified in test serum were normalized to values obtained when cells were incubated with normal control AB serum (Atlanta Biologicals, Norcross, GA, USA). IgG subclass analysis of HLA antibodies was assessed in the same manner using 10 ECP donors and pooled sera from high calculated panel reactive antibody (cPRA) transplant candidates | Southern Biotech |
| Griffiths et al. (132) | Renal/HLA | For each IgG subclass and for every individual bead by using four negative control sera (NC1–4) from healthy, non-sensitized, and HLA antibody-negative cutoff MFI mean NC1–4 + 3 SDs NC1–4. To determine the amount of IgG subclasses, we used the ratio above the corresponding cutoff | Sigma |