We compared dengue virus (DV) isolation rates and tested whether acute primary (P) and acute/probable acute secondary (S/PS) DV infections could be correctly classified serologically when the patients' first serum (S1) samples were obtained 1 to 3 days after the onset of symptoms (AOS). DV envelope/membrane protein-specific immunoglobulin M (IgM) capture and IgG capture enzyme-linked immunosorbent assay (ELISA) titrations (1/log(10) 1.7 to 1 log(10) 6.6 dilutions) were performed on 100 paired S1 and S2 samples from suspected DV infections. The serologically confirmed S/PS infections were divided into six subgroups based on their different IgM and IgG responses. Because of their much greater dynamic ranges, IgG/IgM ELISA titer ratios were more accurate and reliable than IgM/IgG optical density (OD) ratios recorded at a single cutoff dilution for discriminating between P and S/PS infections. However, 62% of these patients' S1 samples were DV IgM and IgG titer negative (<OD(max)/2 titer threshold), and in 35% of the S/PS infections, the patients' S1 and S2 samples were IgM titer negative. The IgM OD values were, however, much higher than those of IgG in the S1 samples of many of these, and the other, S/PS infections. This necessitated using higher (>or=2.60 and <2.60) discriminatory IgM/IgG OD (DOD) ratios on these S1 samples than those published previously to correctly classify the highest percentage of these P and S/PS infections. The DV isolation rate was highest (12/12; 100%) using IgG and IgM titer-negative S1 samples collected 1 day AOS, when 100% of them were correctly classified as P or S/PS infections using these higher DOD ratios.