production of a human recombinant antibody against serotype a candida albicans

after using 3 different generations of antibodies including human and non-human hyperimmune sera, monoclonal antibodies and chimeric antibodies, more recently a newer approach has been developed in which the antibody genes are cloned directly from a patient peripheral b-lymphocytes and expressed in a host like e. coli. in this study the candida albicans serotype a (nctc 3153) mannan was purified using a modified fehling method and used for selection of human recombinant antibody from a c. albicans phage antibody library. after four rounds of affinity selecting (panning), 2 predominant clones were chosen by dna fingerprinting and elisa. a 248 amino acid dna fragment coding for anti-c. albicans mannan scfv was sequenced and cloned in a pbad-topo cloning vector to produce a soluble and phage free antibody. the analysis of antibody sequences by v base index (dnaplot) confirmed the human antibody origin with the vh4 family in v segment of heavy variable chain and vl3 (lambda 3) in j segment of the light variable chain. this antibody fragment was purified using immobilized metal affinity chromatography and inmmunoblotted as a 31kda recombinant protein.