Various biomarkers targeting cell-free DNA (cfDNA) and circulating proteins have been tested for pan-cancer detection. Oncofetal chondroitin sulfate (ofCS), which distinctively modifies proteoglycans (PGs) of most cancer cells and binds specifically to the recombinant Plasmodium falciparum VAR2CSA proteins (rVAR2), is explored for its potential as a plasma biomarker in pan-cancer detection. To quantitate the plasma ofCS/ofCSPGs, we optimized an ELISA using different capture/detection pairs (rVAR2/anti-CD44, -SDC1, and -CSPG4) in a case-control study with six cancer types. We show that the plasma levels of ofCS/ofCSPGs are significantly higher in cancer patients (P values, 1.2 × 10 to 4.4 × 10). Validation studies are performed with two independent cohorts covering 11 malignant tumors. The individuals in the top decile of ofCS-CD44 have more than 27-fold cancer risk (OR = 27.8, 95%CI = 18.8-41.4, P = 2.72 × 10) compared with the lowest 20%. Moreover, the elevated plasma ofCS-CD44 could be detected at the early stage of pan-cancer with strong dose-dependent odds risk prediction.