α2-macroglobulin (α2-M) have crucial clinical significance as a potent biomarker for diabetes nephropathy (DN). There is an increasing demand for rapid detection of α2-M. Herein, a magnetoelastic (ME) biosensor with a layered composite structure is developed for α2-M detection. Based on 3D printing, the basal layer of the biosensor is prepared as a grid structure fabricated by polylactic acid (PLA) doped with NiFe2O4. The conductivity of the biosensor was improved significantly due to the addition of multi-walled carbon nanotubes (MWCNTs). The biological site for capturing α2-M antigen was provided by the carbon quantum dots (CDs) coupled with anti-α-2M (anti-α2-M@CDs) in the nitrocellulose filter (NC) membrane. Meanwhile, the distribution of antibodies on the biosensor surface can be observed more directly due to the fluorescence characteristics of CDs. The biosensor in this work can realize multi-pattern recognition of fluorescent signal and electromagnetic signal. The results show that the limit of detection (LOD) was 0.506 ng/mL in the linear range from 10 ng/mL to 100 µg/mL and the linear equation of fitting curve is: y = 0.21 x - 0.15. The ME biosensors with a simple preparation method have advantages of high sensitivity, good stability and low LOD, showing the great potential for α2-M detection.