Electrophoretic determination of ricin using immunogold silver staining--comparison with simple "protein dot" method.
Studies were performed to establish a sensitive electrophoretic immunodetection system for the highly toxic plant protein, ricin. This has potential criminal application as an agent for causing a delayed death following parenteral administration. The immunodetection system could be used to demonstrate residual traces of the toxin left in certain tissues of the victim's body. Following polyacrylamide gel electrophoresis of ricin added to rat muscle tissue extracts, the gels were electro-blotted onto nitrocellulose paper and ricin bands probed for visualisation by immunostaining. Several immunostaining procedures were investigated in order to select the most sensitive. These included indirect immunoperoxidase, peroxidase-anti-peroxidase (PAP), avidin-biotin complex (ABC) and the immunogold silver staining (IGSS) procedures. The sensitivity of PAP and indirect immunoperoxidase methods were similar at around 50 ng while the ABC technique gave visible staining of 10 ng of electro-blotted ricin. The method with greatest sensitivity was undoubtedly IGSS, which resulted in unequivocal demonstration of 4 ng of ricin. The IGSS-immunoblotting system was considered to readily demonstrate the presence of ricin in muscle tissue from the injection site of dead victims. We compared this system with the very simple method of sample dot staining. Here, samples of ricin were spotted directly onto nitrocellulose. The dots were stained using the IGSS method which was found able to demonstrate less than 10 pg of ricin.
Terazawa
K
K
Griffiths
G D
GD
Leith
A G
AG
Green
M A
MA
D000818:Animals; D004591:Electrophoresis, Polyacrylamide Gel; D009132:Muscles / Q000032:analysis; D051381:Rats; D012276:Ricin / Q000032:analysis* / Q000276:immunology; D012834:Silver; D013194:Staining and Labeling
D012834:Silver; D012276:Ricin