Disease-specific alterations of the cell-free DNA methylation status are frequently found in serum samples and are currently considered to be suitable biomarkers.Candidate markers were identified by bisulfite conversion-based genome-wide methylation screening of lung tissue from lung cancer, fibrotic ILD, and COPD. cfDNA from 400 μl serum (n = 204) served to test the diagnostic performance of these markers. Following methylation-sensitive restriction enzyme digestion and enrichment of methylated DNA via targeted amplification (multiplexed MSRE enrichment), a total of 96 markers were addressed by highly parallel qPCR.Lung cancer was efficiently separated from non-cancer and controls with a sensitivity of 87.8%, (95%CI: 0.67–0.97) and specificity 90.2%, (95%CI: 0.65–0.98). Cancer was distinguished from ILD with a specificity of 88%, (95%CI: 0.57–1), and COPD from cancer with a specificity of 88% (95%CI: 0.64–0.97). Separation of ILD from COPD and controls was possible with a sensitivity of 63.1% (95%CI: 0.4–0.78) and a specificity of 70% (95%CI: 0.54–0.81). The results were confirmed using an independent sample set (n = 46) by use of the four top markers discovered in the study (HOXD10, PAX9, PTPRN2, and STAG3) yielding an AUC of 0.85 (95%CI: 0.72–0.95).This technique was capable of distinguishing interrelated complex pulmonary diseases suggesting that multiplexed MSRE enrichment might be useful for simple and reliable diagnosis of diverse multifactorial disease states.

Saliva based diagnostics is a rapidly evolving field due to the large diagnostic potential and simple sample collection. Currently only few individual molecules were investigated for their diagnostic capabilities in saliva. A systematic comparison of IgG antibody profiles in saliva and plasma is still missing in scientific literature. Our hypothesis is that IgG profiles in plasma and saliva are highly similar for each individual. As a consequence, one could implement practically any plasma based IgG assay (classical serology) as saliva based assay. In other words, the IgG antibodies found in blood are also accessible from saliva. We confirm our hypothesis by comparing IgG reactivities towards protein and peptide antigens. We isolated saliva IgG with high purity and demonstrate that plasma IgG reactivities (classical serology) can be inferred from saliva. As a showcase we perform Hepatitis B virus antibody (plasma-)titer determination from saliva. Additionally we show that plasma and saliva IgG profiles of 20 individuals are highly similar for 256 peptide antigens and match (unsupervised) with high probabilities. Finally, we argue for generalisation to the complete IgG antibody profile. The presented findings could contribute greatly to the development of saliva based diagnostic methods of numerous antibody based tests.

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