Development of an enzyme-linked immunosorbent assay for podophyllotoxin.
A competitive indirect enzyme-linked immunosorbent assay (ciELISA) for podophyllotoxin was developed by using polyclonal antibody, and its suitability for the determination of this analyte in spiked water samples was studied. To avoid antibody production to the linker, the succinoyl-podophyllotoxin (hapten) mimicking the analyte was synthesized and conjugated with the carrier proteins bovine serum albumin (BSA) and ovalbumin (OVA) by mixed anhydride reaction (MAR) and active ester method (AEM). Polyclonal antibodies raised against Hapten-BSA(1) synthesized by MAR and Hapten-BSA(2) by AEM were screened and selected for the ciELISA. One set of antibodies from the rabbit M4440 immunized with Hapten--BSA(1) showed an I(50) value of 2.21 microg/mL with a detection limit of 0.12 microg/mL, and the other set from the rabbit M4469 immunized with Hapten-BSA(2) had an I(50) value of 0.7897 microg/mL with a detection limit of 0.0056 microg/mL. This assay showed the cross-reactivities with the structurally closely related compounds. Recoveries from the podophyllotoxin-fortified tap water in the assay were in the range of 72--115%. A good correlation between podophyllotoxin concentration measured by the ELISA and HPLC (R(2)=0.9924, Y=1.195X-0.257) was obtained from linear regression analysis. These results indicate that the ELISA could be a convenient and supplemental analytical tool for monitoring podophyllotoxin and its analogues in waters without previous extraction or cleanup.
Xu
Dun Ming
DM
R&D Center of Biorational Pesticides, Northwest A & F University, Yangling 712100, China. dunmingxu@163.com
Yu
Xiang Yang
XY
Liu
Ying Qian
YQ
Feng
Jun Tao
JT
Pan
Li Gang
LG
Liu
Xian Jin
XJ
He
Jun
J
Zhang
Xing
X
D000818:Animals; D000906:Antibodies; D000915:Antibody Affinity; D000918:Antibody Specificity; D000941:Antigens; D000970:Antineoplastic Agents / Q000032:analysis*; D003429:Cross Reactions; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D008297:Male; D015394:Molecular Structure; D011034:Podophyllotoxin / Q000032:analysis*; D011817:Rabbits; D012680:Sensitivity and Specificity; D014867:Water / Q000737:chemistry
D000906:Antibodies; D000941:Antigens; D000970:Antineoplastic Agents; D014867:Water; D011034:Podophyllotoxin