Ramos-González M I, Ruiz-Cabello F, Brettar I, Garrido F, Ramos J L
Consejo Superior de Investigaciones Científicas, Departamento de Bioquímica, Vegetal, Granada, Spain.
J Bacteriol. 1992 May;174(9):2978-85. doi: 10.1128/jb.174.9.2978-2985.1992.
Assessment of potential risks involved in the release of genetically engineered microorganisms is facilitated by the availability of monoclonal antibodies (MAbs), a tool potentially able to monitor specific organisms. We raised a bank of MAbs against the soil bacterium Pseudomonas putida 2440, which is a host for modified TOL plasmids and other recombinant plasmids. Three MAbs, 7.3B, 7.4D, and 7.5D, were highly specific and recognized only P. putida bacteria. Furthermore, we developed a semiquantitative dot blot assay that allowed us to detect as few as 100 cells per spot. A 40-kDa cell surface protein was the target for MAbs 7.4D and 7.5D. Detection of the cell antigen depended on the bacterial growth phase and culture medium. The O antigen of lipopolysaccharide seems to be the target for MAb 7.3B, and its in vivo detection was independent of the bacterial growth phase and culture medium. MAb 7.3B was used successfully to track P. putida (pWW0) released in unsterile lake mesocosms.
单克隆抗体（MAb）的可用性有助于评估基因工程微生物释放所涉及的潜在风险，这是一种有可能监测特定生物体的工具。我们制备了一组针对土壤细菌恶臭假单胞菌2440的单克隆抗体，该细菌是修饰的TOL质粒和其他重组质粒的宿主。三种单克隆抗体7.3B、7.4D和7.5D具有高度特异性，仅识别恶臭假单胞菌。此外，我们开发了一种半定量斑点印迹测定法，使我们能够在每个斑点中检测到低至100个细胞。一种40 kDa的细胞表面蛋白是单克隆抗体7.4D和7.5D的靶标。细胞抗原的检测取决于细菌生长阶段和培养基。脂多糖的O抗原似乎是单克隆抗体7.3B的靶标，其体内检测与细菌生长阶段和培养基无关。单克隆抗体7.3B成功用于追踪在未灭菌的湖泊中宇宙释放的恶臭假单胞菌（pWW0）。
