Background: Alzheimer's disease is a progressive neurodegenerative disorder characterized by memory loss and cognitive impairment. The diagnosis of Alzheimer's disease according to symptomatic events is still a puzzling task. Developing a biomarker-based, low-cost, and high-throughput test, readily applicable in clinical laboratories, dramatically impacts the rapid and reliable detection of the disease. Objective: This study aimed to develop an accurate, sensitive, and reliable screening tool for diagnosing Alzheimer's disease, which can significantly reduce the cost and time of existing methods. Methods: We have employed a MALDI-TOF-MS-based methodology combined with a microaffinity chromatography enrichment approach using affinity capture resins to determine serum kappa (κ) and lambda (light chain levels in control and patients with AD. Results: We observed a statistically significant difference in the kappa light chain over lambda light chain (κLC/LC) ratios between patients with AD and controls (% 95 CI: -0.547 to -0.269, p#60;0.001). Our method demonstrated higher sensitivity (100.00%) and specificity (71.43%) for discrimination between AD and controls. Conclusion: We have developed a high-throughput screening test with a novel sample enrichment method for determining κLC/LC ratios associated with AD diagnosis. Following further validation, we believe our test has a potential for clinical laboratories.