Gene expression profiles reflect the biologically diverse activities of cells under specific cell environments. Using the transcriptional response of cultured cells to blood composition, we developed a litmus gene assay to discriminate blood samples reflecting different sample qualities or disease conditions. This cell-based litmus gene assay identified six genes (CCL20, CEMIP, IL1B, IL8, PRG2, PTGS2) as potential biomarkers of plasma quality control and the SPC25 gene as a diagnostic biomarker of Alzheimer's disease (AD). In addition, the SPC25 gene expression level was significantly increased in the cell-based assay using serum samples from patients with mild cognitive impairment (MCI). In conclusion, we demonstrated the effectiveness and potential of a litmus gene assay to detect the orchestrated effects of circulating systemic factors, leading to the successful diagnosis of AD and MCI. This method is broadly applicable to the diagnosis of disease subtypes or patho-physiological stages of complex diseases and tumors.