Wetherall B L, McDonald P J, Johnson A M
Department of Clinical Microbiology, Flinders Medical Centre, Bedford Park, Australia.
J Med Microbiol. 1988 Aug;26(4):257-63. doi: 10.1099/00222615-26-4-257.
A dot-blot hybridisation assay for the detection of Campylobacter pylori was used to compare a 32P-labelled probe with two biotinylated probes and a sulphonated probe. The minimum amount of pure C. pylori DNA that could be detected by the 32P-labelled probe was 100 pg, which corresponded to 5 x 10(4) bacteria. A biotin-labelled DNA (biotin-DNA) probe together with the BluGeneTM detection system produced by Bethesda Research Laboratories (BRL), and a sulphonated probe and ChemiprobeTM detection system (Orgenics) gave similar levels of sensitivity; nylon membranes could be used with both these non-radioactive detection systems. However, a photobiotin-labelled DNA (photobiotin-DNA) probe and detection system produced by Biotechnology Research Enterprises S.A. (BRESA) gave optimum results only with nitrocellulose membranes, and was quantitatively 100 times less sensitive than the other types of probe. The detection systems for the biotin-DNA and photobiotin-DNA probes produced non-specific reactions with crude bacterial blots of heterologous organisms; these non-specific reactions could be removed by treating the dot blots with proteinase K, but not by treatment with RNAase. The sulphonated probe and detection system did not give any reaction with heterologous organism blots.
采用一种用于检测幽门螺杆菌的斑点杂交试验，将一种32P标记探针与两种生物素化探针及一种磺化探针进行比较。32P标记探针能够检测到的纯幽门螺杆菌DNA的最小量为100皮克，这相当于5×10⁴个细菌。一种生物素标记的DNA（生物素-DNA）探针与贝塞斯达研究实验室（BRL）生产的BluGeneTM检测系统，以及一种磺化探针和ChemiprobeTM检测系统（奥根尼克斯公司）具有相似的灵敏度水平；尼龙膜可用于这两种非放射性检测系统。然而，生物技术研究企业股份公司（BRESA）生产的光生物素标记的DNA（光生物素-DNA）探针及检测系统仅在硝酸纤维素膜上能得到最佳结果，并且在定量方面比其他类型的探针灵敏度低100倍。生物素-DNA和光生物素-DNA探针的检测系统与异源生物体的粗细菌印迹产生非特异性反应；这些非特异性反应可通过用蛋白酶K处理斑点印迹来消除，但不能通过用RNA酶处理来消除。磺化探针及检测系统与异源生物体印迹未产生任何反应。
