Development of a novel antibody probe useful for domoic acid detection.
The generation of monoclonal antibody (mAb) against marine toxins can serve as a valuable probe to detect this kind of compounds by immunological methods. However, traditional approaches to mAb generation usually need a comparative large quantity of standard substance (more than 400 microg mouse(-1)), and a comparative long immunization period (more than 6 weeks). Here we report a simple, inexpensive and fast protocol for the generation of monoclonal antibody probe specific for domoic acid (DA). In the method, lymph node cells were harvested from the Balb/C mice of hind footpad injection and fused with murine myeloma cells SP2/0 for hybridoma generation. This method for the preparation of mAb for DA has two main advantages: (a) there is no need for large-scale expensive antigen (only 70 microg antigen for one mouse); (b) immunization protocol can be accomplished within 16 days. Some characteristics of the mAb were studied and a direct competitive ELISA for the detection of DA using the mAb as a probe was developed. The detection limit was 0.41 ng well(-1) in phosphate buffered saline (PBS) and 0.53 ng well(-1) in blue mussel Mytilus edulis. The recoveries of DA from mussel and PBS buffer were from 94.8% to 105.1% and from 96.2% to 103.7%, respectively. Thus, the newly developed direct competitive ELISA using the mAb appears to be a reliable and useful method for monitoring of DA in shellfish (228 words).
Zhou
Yu
Y
Key Laboratory of Zoonosis Research, Ministry of Education, Institute of Zoonosis, Jilin University, Changchun 130062, PR China. zhouyu69@sina.com
Zhang
Yuan-Yuan
YY
Shen
Qing-Feng
QF
Lu
Shi-Ying
SY
Ren
Hong-Lin
HL
Li
Yan-Song
YS
Liu
Zeng-Shan
ZS
Pan
Feng-Guang
FG
Meng
Xian-Mei
XM
Zhang
Jun-Hui
JH
D000818:Animals; D000911:Antibodies, Monoclonal; D045744:Cell Line, Tumor; D004587:Electrophoresis, Agar Gel; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D005260:Female; D006825:Hybridomas; D007114:Immunization / Q000379:methods; D007608:Kainic Acid / Q000031:analogs & derivatives* / Q000032:analysis / Q000276:immunology; D008297:Male; D008387:Marine Toxins / Q000032:analysis* / Q000276:immunology*; D051379:Mice; D008807:Mice, Inbred BALB C; D049880:Mytilus edulis / Q000737:chemistry
D000911:Antibodies, Monoclonal; D008387:Marine Toxins; C012301:domoic acid; D007608:Kainic Acid