We have developed immunopillar devices for rapid and easy-to-use immunoassay with pM-fM detection sensitivity, but long total assay time (sample-in-answer-out) and the compatibility with a portable detection system have still remained as major problems toward the point-of-care testing (POCT). We report here next generation immunopillar devices and portable detection system suitable for them. Thin structure of those devices allowed us to wash nonspecifically bound antigens and fluorescent-labeled secondary antibodies in a minute, resulting in the dramatic reduction in the total assay time. In addition, by using the portable detection system, we gained the concentrations of C-reactive protein in human sera while preserving pM detection sensitivity. The total assay time was 20 minutes per sample. Our immunoassay system possesses the potential for use in POCT.