ISOLATION OF IGG ANTIBODY FV-DNA FROM VARIOUS MOUSE AND RAT HYBRIDOMACELL-LINES USING THE POLYMERASE CHAIN-REACTION WITH A SIMPLE SET OF PRIMERS

To facilitate the isolation of IgG antibody FV-DNA sequences from hybr idoma cell lines, we have established a polymerase chain reaction (PCR ) procedure requiring only a small number of primers. The sense primer s homologous to DNA coding for the first framework sequences were desi gned to hybridize to all the known antibody sequences under conditions that permit a high number of mismatches. The antisense primers were h omologous to DNA coding for the beginning of the constant regions of t he gamma and kappa chains. Restriction sites introduced by the primers enable the DNA to be cloned into bacterial expression vectors. Only t hree sense V-H primers and two sense V-L primers paired with one backw ard primer for the heavy and light chains, respectively, were necessar y for theification of Fv-DNA from a total of 17 rodent cell lines that we have so far worked with. These consisted of 12 mouse cell lines an d five rat cell lines. This procedure will therefore probably be suffi cient to isolate the Fv-DNA from most mouse cell lines and possibly al so from most rat cell lines.