The serum anti-Ancylostoma duodenale immunoglobulin (Ig) G4 antibody response to fraction III of the partially purified excretory secretory antigen of adult worm (Ad III ESA) was studied. The work included 60 patients with A. duodenale infection (GI), 40 patients with other parasitic infections (GII) and 30 apparently healthy parasite-free controls (GIII). Level of serum specific IgG4 was measured by indirect enzyme linked immunosorbent assay and compared with serum specific IgG, IgG 1, 2 & 3 subclass antibodies. Patients of GI had gastro-intestinal manifestations and symptoms suggestive of anaemia, and by investigations they had anaemia in 31.7% & eosinophilia in 100%. Measuring the intensity of A. duodenale infection, quantified as fecal egg counts, in patients of GI revealed that 60%, 30% & 10% had light, moderate, and heavy infections, respectively. The serum anti-Ad III ESA IgG & IgG 1-4 subclass antibodies were significantly elevated (P < 0.001) in patients of GI compared with GIII. Serum specific IgG4 was expressed in 100% of patients of GI at a significantly highly elevated level than IgGI (P < 0.01), IgG2 & IgG3 (P < 0.001). Specific IgG1 was expressed in 88.3% of patients of GI at a significantly elevated level (P < 0.001) than IgG2 & IgG3 which were expressed in 31.7% & 38.3%, respectively and elevated to a moderate extent. Serum specific IgG4 showed a 1.0, 1.1, 3.1 & 2.6-fold increase in detection rate of positive cases than IgG, IgG1, IgG2 & IgG3, respectively. The highest ability to differentiate between infected and healthy subjects was by serum specific IgG4 recording a discrimination coefficient of 9.4, while IgG, IgG1, IgG2 & IgG3 recorded 5.2, 6.3, 3.2 & 3.4, respectively. Serum specific IgG4 showed a significant positive correlation (r = 0.41, P < 0.001) with the intensity of A. duodenale infection, as was demonstrated by IgG & IgG3 (P < 0.01 & P < 0.05, respectively). Detection of serum anti-Ad III ESA IgG4 antibody recorded a 100% sensitivity that was significantly higher (P < 0.001) than IgG1, IgG2 & IgG3, but insignificantly different (P > 0.05) from IgG. Finally, serum specific IgG4 recorded a 100% specificity that was significantly higher than IgG, IgG2, IgG3 (P < 0.01) & IgG1 (P < 0.05). They showed cross-reactions with ascariasis, lymphatic filariasis and strongyloidiasis. The results are discussed.