Objective.Serum 14-3-3η is a novel joint-derived proinflammatory mediator implicated in the pathogenesis of rheumatoid arthritis (RA). In our study, we assessed the diagnostic utility of 14-3-3η and its association with standard clinical and serological measures.Methods.A quantitative ELISA was used to assess 14-3-3η levels. Early (n = 99) and established patients with RA (n = 135) were compared to all controls (n = 385), including healthy subjects (n = 189). The sensitivity, specificity, positive and negative predictive values of 14-3-3η, and the likelihood ratios (LR) for RA were determined through receiver-operator curve analysis. The incremental value of adding 14-3-3η to anticitrullinated protein antibody (ACPA) and rheumatoid factor (RF) in diagnosing early and established RA was assessed.Results.Serum 14-3-3η differentiated established patients with RA from healthy individuals and all controls (p < 0.0001). A serum 14-3-3η cutoff of ≥ 0.19 ng/ml delivered a sensitivity and specificity of 77% and 93%, respectively, with corresponding LR positivity of 10.4. At this cutoff in early RA, 64% of patients with early RA were positive for 14-3-3η, with a corresponding specificity of 93% (LR+ of 8.6), while 59% and 57% were positive for ACPA or RF, respectively. When ACPA, RF, and 14-3-3η positivity were used in combination, 77 of the 99 patients (78%) with early RA were positive for any 1 of the 3 markers. Serum 14-3-3η did not correlate with C-reactive protein, erythrocyte sedimentation rate, or Disease Activity Score, but patients who were 14-3-3η-positive had significantly worse disease.Conclusion.Serum 14-3-3η is a novel RA mechanistic marker that is highly specific, associated with worse disease, and complements current markers, enabling a more accurate diagnosis of RA.

Fecal neopterin concentration is increased in patients with clinically active or inactive Crohn's disease and in patients with clinically active ulcerative colitis when compared with controls, and therefore represents a new biomarker for disease activity.

Laboratory diagnosis of von Willebrand disease type 2N (VWD2N) is based on costly mutation analysis or in vitro measurement of the ability of plasma von Willebrand factor (VWF) to bind exogenous factor VIII (FVIII); however, the VWF-FVIII binding activity assay is complex and not widely used. Our aim was to assess the utility of the in-house VWF-FVIII binding assay in the investigation of patients with suspected VWD2N. A previously described ELISA-based FVIII binding method was simplified and adapted for the clinical laboratory use by optimizing incubation time, reagent concentrations and assay standardization. The assay was validated using samples from eight individuals with known homozygous or heterozygous VWD2N mutations, and 100 healthy adults. An additional 392 patient samples were tested, including 314 with FVIII activity <50% of normal and 78 received for routine VWF-FVIII binding activity testing. Intra- and inter-assay variations were less than 10% and 17%, respectively, and the limit of quantification was estimated as 0.12. The reference range for healthy adults was 0.73-1.42. VWF:FVIII binding activity was consistent with the genotype in subjects with available genetic data, being low in three individuals with homozygous mutation (<0.12) and intermediate in five heterozygous individuals (0.44-0.61). Screening of the 392 clinical samples identified reduced VWF:FVIII binding in 19 subjects. This assay provides accurate measurement of VWF:FVIII binding activity and successfully identifies homozygous VWD2N patients and heterozygous carriers. Use of this ELISA-based assay may help avoid the need for mutation analysis in patients with unexplained low FVIII activity.

Summary.  The evaluation of a prolonged aPTT often includes Lupus Anticoagulant, Antiphospholipid Antibodies, and Factor VIII (FVIII) inhibitors. We have noticed that patient samples positive for lupus antibody (LA) are frequently also positive for FVIII IgG antibodies in an enzyme‐linked immunosorbent assay (ELISA), indicating the need for follow‐up testing with a more labour‐intensive functional assay for FVIII inhibition. This study evaluates the potential for a FVIII IgG ELISA to yield false‐positive results in patient samples positive for LA or other antiphospholipid antibodies. A total of 289 residual de‐identified patient samples positive for LA (n = 143), anti‐cardiolipin IgG (n = 84), or beta2‐glycoprotein antibody (n = 62) were tested for FVIII IgG using a commercial ELISA. Samples with positive FVIII IgG ELISA results were further tested for FVIII activity using a clot‐based FVIII inhibitor assay. The FVIII IgG ELISA yielded positive results in 39 (13%) of the samples tested, including 13/143 (13%) LA‐positive, 15/85 (18%) aCL IgG‐positive and 6/62 (10%) β2‐glycoprotein IgG‐positive samples. The clot‐based FVIII inhibitor assay yielded negative results in all 39 FVIII IgG‐positive specimens tested, indicating discrepancy with the FVIII IgG ELISA results. Patient specimens positive for LA, aCL IgG, or β2‐glycoprotein IgG may yield false‐positive results for FVIII antibodies. Caution is warranted in interpreting FVIII antibody results in these cases.

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