MOLECULAR BIOLOGICAL TECHNIQUES FOR SUBSPECIES IDENTIFICATION: IMMUNOLOGICAL TECHNIQUES A COMPARISON

摘要

Several immunological techniques such as ELISA or western blotting are available that offer means of relatively easy and rapid identification of microbial agents at realistic limits of detection, that is, in the nanogram to femtogram ranges. Other techniques that are somewhat more difficult to perform such as surface plasmon resonance spectroscopy nevertheless in controlled experiments permit detection of subtle changes in antigen epitope topology that can provide valuable information about the character of the microorganism, for example, if it is a mutant. Such methods can be used as a first indication of variance by a particular strain of microorganism, but the last proof has to be obtained by genetic analysis, for example information provided by PCR or nucleotide sequencing. In order to detect strain variants by immunological methods, antigenic variation must reflect strain variation, and there must be a panel of antibodies available with defined binding properties for all different possible antigenic variants. This is a formidable criterion that has to be met, but one approach might be to develop antibodies through molecular biological techniques that bind to conserved or generic determinants of the antigen in question, and follow up this assay with another using antibodies recognizing strain differences. It would also be useful to have a panel of antibodies available that recognize different antigenic structures on a microorganism, with the restriction that those antigenic structures are characteristic only for that particular microorganism. In other situations, immunological typing is less useful. This is illustrated by the group of enteropathogenic Escherichia coli bacteria. While some of these pathogenic organisms are restricted to certain serotypes, the serotype is not a definitive indication that the organism is pathogenic. In this case, molecular biological analysis techniques are more informative. Usually, a combination of immunological and molecular biological techniques would be useful. The immunological techniques are easily performed and could in well defined cases be used as a screening procedure that could be followed up with a genetic based analysis. It should, however, be recalled that for the detection of toxins apart from the organisms that produce them, genetic analyses are not applicable.