Gibson A M, Modi N K, Roberts T A, Hambleton P, Melling J
Agricultural and Food Research Council, Institute of Food Research, Bristol Laboratory, Langford, UK.
J Appl Bacteriol. 1988 Apr;64(4):285-91. doi: 10.1111/j.1365-2672.1988.tb01873.x.
A monoclonal antibody-based amplified ELISA method for detecting Clostridium botulinum type B toxin was evaluated for its ability to detect the toxin in the supernatant fluid of pure cultures and after growth from Cl. botulinum spores inoculated into pork slurries. Slurries containing NaCl (1.5-4.5% w/v) and polyphosphate (0.3% w/v) were either unheated or heated 80 degrees C/5 min followed by 70 degrees C/2 h before incubation at 15 degrees, 20 degrees or 27 degrees C. Presence of specific toxin was confirmed by mouse bioassay and results were compared with those of the amplified ELISA method. A total of 48 strains, consisting of 38 Cl. botulinum and 10 Cl. sporogenes (putrefactive anaerobes), and 140 slurry samples were tested. Cultures of eight out of nine strains of type B Cl botulinum and 73 of 101 slurry samples containing type B toxin were positive by ELISA; the remaining 28 slurry samples contained type B toxin at levels below or close to the detection limit (20 LD50/ml) of the type B ELISA. No false-positive reactions occurred with Cl. botulinum types A, C, D, E or F, or with the 10 strains of Cl. sporogenes. Toxin produced by one strain of Cl. botulinum type B (NCTC 3807) was not detected by this single monoclonal antibody-based amplified ELISA. With a mixture of two monoclonal antibodies, however, the toxin from NCTC 3807 could be detected without reducing the sensitivity of the ELISA.
评估了一种基于单克隆抗体的扩增酶联免疫吸附测定（ELISA）方法，用于检测B型肉毒梭菌毒素，该方法用于检测纯培养物上清液以及接种到猪肉匀浆中的肉毒梭菌孢子生长后的毒素。含有氯化钠（1.5 - 4.5% w/v）和多聚磷酸盐（0.3% w/v）的匀浆，要么不加热，要么在80℃加热5分钟，然后在70℃加热2小时，之后在15℃、20℃或27℃下培养。通过小鼠生物测定法确认特定毒素的存在，并将结果与扩增ELISA方法的结果进行比较。共测试了48株菌株，包括38株肉毒梭菌和10株生孢梭菌（腐败厌氧菌），以及140个匀浆样品。9株B型肉毒梭菌中的8株培养物以及101个含B型毒素的匀浆样品中的73个通过ELISA呈阳性；其余28个匀浆样品中B型毒素的含量低于或接近B型ELISA的检测限（20 LD50/ml）。A、C、D、E或F型肉毒梭菌以及10株生孢梭菌均未出现假阳性反应。该基于单克隆抗体的扩增ELISA未检测到一株B型肉毒梭菌（NCTC 3807）产生的毒素。然而，使用两种单克隆抗体的混合物，可以检测到NCTC 3807产生的毒素，且不会降低ELISA的灵敏度。
