This paper describes enzyme-linked immunosorbent assays (ELISAs) utilizing microfluidic thread/paper-based analytical devices (TPAD), microfluidic fabric-based analytical devices (FAD), and microfluidic thread-based analytical devices (TAD). Here, the quantitative detection of biotinylated goat anti-mouse IgG (system one) and rabbit IgG (system two) antibodies via colorimetric analysis is detailed. In both systems, antibody is spotted on the detection site and subjected to a series of washes, addition of streptavidin-alkaline phosphatase (Strep-ALP) (system 1) or alkaline phosphatase (ALP)-conjugated secondary antibody (system 2), and colorimetric substrate. The devices are scanned and analyzed yielding a correlation between inverse yellow (or purple) intensity. For system one, a linear range of detection at low concentrations of streptavidin-alkaline phosphatase (Strep-ALP) was observed befire the enzyme reached a V-max. At higher concentrations of Strep-ALP, saturation is achieved for both the TPAD and FAD devices. For system two, the IC50 values obtained for the non-trifurcated and trifurcated TADs were determined to be 180.2fmol/zone and 133.8fmol/zone, respectively. The IC50 value was demonstrated to be 1034fmol/zone and 208.6fmol/zone for the TPADs and FADs, respectively. For all devices the lowest concentration of Strep-ALP or rabbit IgG used in the assay was 3.75 x 10(-4)mg/mL and 0.7fmol/zone, respectively. The development of this technology should further facilitate the use of these platforms for ELISA to detect and quantitate antibodies.