Detection of ricin and other ribosome-inactivating proteins by an immuno-polymerase chain reaction assay.
Ribosome-inactivating proteins (RIPs) are plant proteins with enzymatic activity, classified as type 1 (single chain) or type 2 (two chains). They are identified as rRNA N-glycosidases (EC 3.2.2.22) and cause an irreversible inhibition of protein synthesis. Among type 2 RIPs, there are potent toxins (ricin is the best known) that are considered as potential biological weapons. The development of a fast and sensitive method for the detection of biological agents is an important tool to prevent or deal with the consequences of intoxication. In this article, we describe a very sensitive immuno-polymerase chain reaction (IPCR) assay for the detection of RIPs-a type 1 RIP (dianthin) and a type 2 RIP (ricin)-that combines the specificity of immunological analysis with the exponential amplification of PCR. The limit of detection (LOD) of the technique was compared with the LODs of the conventional immunological methods enzyme-linked immunosorbent assay (ELISA) and fluorescent immunosorbent assay (FIA). The LOD of IPCR was more than 1 million times lower than that of ELISA, allowing the detection of 10 fg/ml of dianthin and ricin. The possibility to detect ricin in human serum was also investigated, and a similar sensitivity was observed (10 fg/ml). IPCR appears to be the most sensitive method for the detection of ricin and other RIPs.
Lubelli
Chiara
C
Department of Experimental Pathology, University of Bologna, via S. Giacomo 14, I-40126 Bologna, Italy.
Chatgilialoglu
Alexandros
A
Bolognesi
Andrea
A
Strocchi
Paola
P
Colombatti
Marco
M
Stirpe
Fiorenzo
F
D019921:Biotinylation; D004247:DNA / Q000032:analysis / Q000737:chemistry / Q000235:genetics; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D015200:Fluoroimmunoassay / Q000379:methods; D006801:Humans; D008962:Models, Theoretical; D009699:N-Glycosyl Hydrolases / Q000032:analysis* / Q000235:genetics; D010940:Plant Proteins / Q000032:analysis* / Q000235:genetics; D016133:Polymerase Chain Reaction / Q000379:methods*; D015203:Reproducibility of Results; D054789:Ribosome Inactivating Proteins, Type 1; D012276:Ricin / Q000032:analysis / Q000235:genetics
D010940:Plant Proteins; D054789:Ribosome Inactivating Proteins, Type 1; D004247:DNA; D012276:Ricin; D009699:N-Glycosyl Hydrolases