tailieunhanh - Development of a lateral flow immunoassay strip for rapid detection of rotavirus in fecal samples

In this study, we present the development of a lateral flow immunoassay for detection of rotavirus using polyclonal is the most common cause of severe, dehydrating diarrhea in children worldwide. | Protein A can bind specifically to Fc regions of many mammalian immunoglobulins and commonly used as affinity absorbents to purify antibodies from serum. In present study, protein A - sepharose 4B conjugate was used for affinity purification of anti-rotavirus IgGs from sera of rabbit and guinea pig. The result of rabbit IgG purification shows that there were only two separate peaks on the chromatographic chart with retention times of and min, respectively (fig. 1A). The first peak was present just after loading sample ( min) indicating that proteins in this fraction were unbound proteins. The second peak was present just after adding the low pH glycine solution (pH ) indicating that proteins in this fraction were protein A specific binding IgGs. Under low pH condition the weak linkages between protein A and IgGs will be broken to release IgGs in eluent fraction. The protein pattern of eluent fraction was analyzed by SDS-PAGE and showed the presence of two protein bands with size of approximately 50 and 25 kDa (fig. 1, lane 3). These bands were the IgG heavy chain and light chain, respectively. Observed results demonstrated that IgG was purified from serum using protein A-sepharose gel. After PBS pH buffer exchange using Amicon Ultra-4 30K, the amount of purified IgG was determined as mg/ml serum by bradford assay.