The non-invasive approach of finding biomarkers in peripheral blood of cancer patients makes it useful for clinical application and cancer screening. The aim of the study was to explore the clinical utility of alpha-crystallin antibodies as markers for diagnosis of non-small cell lung cancer (NSCLC) and screening among high-risk groups. Alpha-crystallin antibodies were detected with enzyme-linked immunosorbent assay (ELISA) in 51 NSCLC patients, 38 high-risk chronic obstructive pulmonary disease (COPD) patients and 52 age and sex matched healthy volunteers. Alpha-crystallin IgG antibodies differed significantly between the groups of cancer patients and the healthy volunteers (P<0.001). A cut-off value of 0.317 discerned NSCLC patients with sensitivity 62%, and specificity 72% among the control group. The assay was effective in distinguishing the patients with and without lymphogenic metastatic spread of the disease (P=0.045): sensitivity 60%, and specificity 70%. The clinical significance of this marker has a modest implication in lung cancer diagnosis and screening in high-risk groups. Its importance as a prognostic marker or a marker of disease recurrence and lymph node micrometastasis should be further explored.