husplint_10 independent pcr reactions to amplify v gene segments using all possible combinations within a primer set for husplint_09 see table i for husplint_10 see table ii the primer sequences which in theory encompass the entire repertoire of human antibody genes were obtained from imgt gene db giudicelli et al 2005 stud health technol inform 116 3 8 and modified according to previously published protocols sblattero and bradbury 1998 immunotechnology 3 271 278 marks et al 1991 eur j immunol 21 985 991 orlandi et al 1992 biotechnology 24 527 531 in this method the individual rearranged heavy and light chain variable regions are amplified separately and are linked through a series of overlapping polymerase chain reaction pcr steps to give the final scfv products that are used for cloning visintin et al 2004 j immunol methods 290 135 153 the pcr reactions for husplint_9 table i included seven vh forward primers paired with four vh reverse primers which generated a total of twenty eight reactions whereas four v u03ba forward primers paired with four reverse primers generated a total of sixteen reactions and nine v u03bb forward primers paired with two v u03bb reverse primers generated a total of eighteen reactions the pcr reactions for husplint_10 table ii included six vh forward primers paired with four vh reverse primers.