Bosompem K M, Assoku R K, Nantulya V M
Noguchi Memorial Institute for Medical Research, Legon, Accra, Ghana.
Parasitology. 1996 Jan;112 ( Pt 1):59-66. doi: 10.1017/s0031182000065070.
A sensitive and specific nitrocellulose (NC) membrane-based dot-ELISA, utilizing a panel of monoclonal antibodies (mAbs), was developed for differentiation between in vitro-derived procyclic forms of Trypanosoma brucei, T. congolense and T. simiae, and epimastigotes of T. vivax. Trypanosomes in suspension were applied onto NC membrane in dots and probed with unlabelled trypanosome species-specific mAbs. Bound mAb was revealed by enzyme labelled anti-mouse IgG and precipitable chromogenic substrate. The assay detected the aforementioned trypanosome species in both single and artificially mixed preparations. Ten T. brucei, 4 T. vivax, 7 T. congolense and 3 T. simiae procyclic stocks and clones from different geographical areas were tested and identified using the specific mAbs in the dot-ELISA which had a specificity of 100%. Some of the T. brucei, T. congolense and Nannomonas-specific mAbs could detect as few as 10 trypanosomes/dot, whilst 1 T. vivax mAb was able to detect a minimum of 100 trypanosomes/dot in monospecies preparations. A concentration of 1 x 10(4) trypanosomes/microliters/dot was eventually determined as ideal for testing in the dot-ELISA. Antigen dots stored at 4 degrees C under desiccated conditions did not show any loss in activity for up to 90 days. However, when stored under similar conditions at room temperature (17-26 degrees C), the T. congolense-specific antigen remained unaffected up to 60 days, and then showed decreased activity when tested on day 90.
利用一组单克隆抗体（mAb），开发了一种基于硝酸纤维素（NC）膜的灵敏且特异的斑点酶联免疫吸附测定法（dot-ELISA），用于区分体外培养的布氏锥虫、刚果锥虫和西氏锥虫的前循环型，以及间日锥虫的上鞭毛体。将悬浮液中的锥虫点样于NC膜上，并用未标记的锥虫种特异性单克隆抗体进行检测。通过酶标记的抗小鼠IgG和可沉淀显色底物来显示结合的单克隆抗体。该测定法可在单一和人工混合制剂中检测上述锥虫种类。使用dot-ELISA中的特异性单克隆抗体对来自不同地理区域的10株布氏锥虫、4株间日锥虫、7株刚果锥虫和3株西氏锥虫的前循环型虫株和克隆进行了检测和鉴定，其特异性为100%。一些布氏锥虫、刚果锥虫和纳诺莫纳属特异性单克隆抗体在单一种类制剂中可检测到低至每点10个锥虫，而1种间日锥虫单克隆抗体在单一种类制剂中至少可检测到每点100个锥虫。最终确定每微升每点1×10⁴个锥虫的浓度为在dot-ELISA中进行检测的理想浓度。在干燥条件下于4℃储存的抗原点在长达90天内未显示任何活性损失。然而，当在室温（17 - 26℃）下在类似条件下储存时，刚果锥虫特异性抗原在60天内保持不受影响，然后在第90天进行检测时显示活性下降。
