Monoclonal antibodies to 2,4-dichlorophenol hydroxylase as probes for the 2,4-D-degradative phenotype

Two different monoclonal antibodies (MAb) were raised against 2,4-dichlorop henol hydroxylase (DCP-hydroxylase) of Ralstonia eutropha JMP134 (pJP4), th e second enzyme in the 2,4-D-degradative pathway of this bacterium. The uti lity of these antibodies in detecting and characterizing 2,4-D-degrading so il bacteria was investigated. One MAb (F6) reacted with DCP-hydroxylase fro m 27 out of 36 strains tested, while the other (MAI, C3) reacted with only 17 isolates. When used with the colony blot technique, MAb F6 was useful fo r detecting cross-reacting strains on plates of pure cultures or of mixture s containing nondegraders even when 2,4-D degraders were outnumbered 60 to 1. 2,4-D-degrading strains could also be detected from plates spread with e nrichment cultures but not from primary isolation plates spread from soil d ilutions, presumably because the ratio of degraders to nondegraders was too low. Colonies of some strains that were very distantly related genetically but produced functionally similar DCP-hydroxlase enzymes, were detected by MAb F6. This result suggests that MAbs could be useful for detecting funct ionally similar proteins expressed from tfdB analogs, even in the absence o f detectable DNA homology between the genes encoding them.