Eintrag weiter verarbeiten
Rapid Immunoassays for Detection of UV-Induced Cyclobutane Pyrimidine Dimers in Whole Bacterial Cells
Gespeichert in:
| Zeitschriftentitel: | Applied and Environmental Microbiology |
|---|---|
| Personen und Körperschaften: | , |
| In: | Applied and Environmental Microbiology, 68, 2002, 5, S. 2542-2549 |
| Format: | E-Article |
| Sprache: | Englisch |
| veröffentlicht: |
American Society for Microbiology
|
| Schlagwörter: |
| Zusammenfassung: | <jats:title>ABSTRACT</jats:title> <jats:p> Immunoassays were developed to measure DNA damage retained by UV-irradiated whole bacterial cells. Active <jats:italic>Mycobacterium parafortuitum</jats:italic> and <jats:italic>Serratia marcescens</jats:italic> cells were fixed and incubated with cyclobutane pyrimidine dimer-binding antibodies after being exposed to known UV doses (254 nm). When both fluorescent (Alexa Fluor 488) and radiolabeled ( <jats:sup>125</jats:sup> I) secondary antibodies were used as reporters, indirect whole-cell assays were sensitive enough to measure intracellular UV photoproducts in <jats:italic>M. parafortuitum</jats:italic> and <jats:italic>S. marcescens</jats:italic> cells as well as photoenzymatic repair responses in <jats:italic>S. marcescens</jats:italic> cells. For the same UV dose, fluorescent DNA photoproduct detection limits in whole-cell assays (immunofluorescent microscopy) were similar to those in fluorescent assays performed on membrane-bound DNA extracts (immunoslot blot). With either fluorescent or radiolabeled reporters, the intracellular cyclobutane pyrimidine dimer content of UV-irradiated whole bacterial cells could be reliably quantified after undergoing a <0.5-order-of-magnitude decrease in culturability. Immunofluorescent microscopy results showed that photoenzymatic repair competence is not uniformly distributed among exponential-growth UV-irradiated pure cultures. </jats:p> |
|---|---|
| Umfang: | 2542-2549 |
| ISSN: |
0099-2240
1098-5336 |
| DOI: | 10.1128/aem.68.5.2542-2549.2002 |