Quantitation of Acinetobacter calcoaceticus in mixed bacterial cultures by an enzyme immunoassay

: An enzyme-linked immunosorbent assay using polyclonal antibodies from rabbits has been developed for quantification of Acinetobacter calcoaceticus. Bacteria were added to the wells of a microtiter plate coated with anti-Acinetobacter immunoglobulin. For detecting bound cells the peroxidase-labelled immunoglobulin fraction was used. Over a distinct range there is a linear correlation between bound bacteria and measured absorbance allowing a quantification of bacteria in an order from 10(7) to 10(8) per milliliter. The specificity of the assay was evaluated by the heterologous bacteria Pseudomonas putida, Pseudomonas aeruginosa, Proteus vulgaris, Escherichia coli and Citrobacter freundii. Only a minimal cross-reactivity was observed. Within a certain range of error it is possible to quantitate Acinetobacter calcoaceticus in mixtures with one or several other bacterial species. Mixed with bacteria of one other species the differences to the value for Acinetobacter calcoaceticus alone do not exceed +/- 10% with a tendency to lower values. Mixed with several other species only negative differences up to -15% were obtained. Treatment of Acinetobacter calcoaceticus with 0.5% formaldehyde results in a loss of reactivity up to 15%. In conclusion, the enzyme-linked immunosorbent assay is a useful method for quantitating bacteria not only with respect to the high sensitivity, specificity and good reproducibility but also for the minimal technical equipment and the short assay time.