Introduction: Type 1 interferons are critical in initial anti-viral responses. Increased type 1 interferon signature (IFNsg), initially described for systemic autoinflammatory disorders (SAID) including Aicardi Goutières (AGS) and proteasome-associated autoinflammatory disorders (PRAAS), is also used as a biomarker for other inflammatory conditions including systemic lupus and dermatomyositis. Effectively measuring the IFNsg is important to understand and properly manage these conditions. However, this test is not widely standardized. Moreover, it is expensive to run on research setting and clinically unavailable. We aimed to improve and optimize the measurement of IFNsg in SAID and dermatomyositis patients by comparing interferon gene expression in whole blood (WB) and peripheral blood mononuclear cells (PBMC). Methodology: IFNsg was measured using RT-qPCR, as previously reported, in samples obtained at the same time point from both, PBMC and WB from 10 patients (9 SAID and 1 dermatomyositis) and 6 controls. We report the Interferon stimulated genes score (ISGs) and fold change for each individual gene. Results: While ISGs from WB were comparable between controls, the fold change for each gene differed between PBMC and WB; SIGLEC1 expression was higher, while IFI27 and IFIT1 were significantly lower in control PBMC. Unlike controls, the comparison of ISGs in PBMC and WB of patients showed significant differences, which were also observed at an individual gene level. These differences were heterogeneous among patients with distinct diagnoses, were mainly observed for MX1, RSAD2, ISG15, IFI27, IFIT1, and OAS1. Conclusion: IFNsg in PBMC and WB varies widely among patients owing to the differential expression of these genes in immune and some non-immune cells. Evaluating IFNsg in WB may disturb the measurement due to type 1 IFNs derived from other cell types, masking immune cell expression patterns in PBMCs. The IFNsg analysis in PBMC may provide more specific information about immune cells relevant to IEI.