Department of Biology, Faculty of Science, Srinakharinwirot University, Bangkok 10110, Thailand.
Biosens Bioelectron. 2013 Apr 15;42:229-35. doi: 10.1016/j.bios.2012.10.011. Epub 2012 Oct 13.
A strip test for the detection of Vibrio cholerae O139 was developed using two monoclonal antibodies (MAbs), namely VC-273 and VC-812, which specifically bind to the lipopolysaccharide and capsular polysaccharide of V. cholerae O139. The MAb VC-273 gold nanoparticle conjugate was sprayed onto a glass fiber pad that was placed adjacent to a sample chamber. MAb VC-812 and the goat anti-mouse immunoglobulin G (GAM) antibody were sprayed onto a nitrocellulose membrane in strips at positions designated as T and C, respectively. The test strips were assessed for their ability to directly detect V. cholerae O139 using samples dispersed in application buffer, and a 100 μL aliquot of sample was applied to the sample chamber. The results were observable within 20 min after application of the sample. In samples containing V. cholerae O139, the antigen was bound to the colloidal gold-conjugated MAb to form an antibody-antigen complex. This complex was captured by the MAbs at the T test line, resulting in the appearance of a reddish-purple band at the T position. The sensitivity of the test was determined to be 10⁴ cfu mL⁻¹. Direct detection of V. cholerae O139 in various fresh seafood samples could be accomplished with similar sensitivities. The detection limit was substantially improved to 1 cfu mL⁻¹ of the original bacterial content after pre-incubation of the sample in alkaline peptone water for 12 h. The V. cholerae strip test provides several advantages over other methods, including the speed and simplicity of use because there is no requirement for sophisticated equipment.
一种用于检测霍乱弧菌 O139 的条带检测法已经开发出来，该方法使用了两种单克隆抗体（MAbs），即 VC-273 和 VC-812，它们特异性地结合霍乱弧菌 O139 的脂多糖和荚膜多糖。MAb VC-273 金纳米颗粒缀合物被喷涂到放置在样品室旁边的玻璃纤维垫上。MAb VC-812 和山羊抗小鼠免疫球蛋白 G（GAM）抗体分别以条带形式喷涂到硝酸纤维素膜上，位置分别标记为 T 和 C。评估了这些测试条带直接检测霍乱弧菌 O139 的能力，使用分散在应用缓冲液中的样品进行测试，将 100 μL 样品加样到样品室中。加样后 20 分钟内即可观察到结果。在含有霍乱弧菌 O139 的样品中，抗原与胶体金缀合的 MAb 结合形成抗体-抗原复合物。该复合物被 MAbs 在 T 测试线上捕获，导致 T 位置出现红紫色带。该测试的灵敏度被确定为 10⁴ cfu mL⁻¹。用类似的灵敏度可以直接检测各种新鲜海鲜样品中的霍乱弧菌 O139。通过将样品在碱性蛋白胨水中孵育 12 小时，将原始细菌含量的检测限提高到 1 cfu mL⁻¹。与其他方法相比，霍乱弧菌条带检测法具有许多优势，包括速度和使用简便性，因为不需要复杂的设备。
