Indian Journal of Public Health Research & Development

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A Pilot Project to Develop in Vivo Detection of Ricin through Fluorescence Sandwich ELISA Technique


Affiliations
1 Department of Biochemistry Saraswathi Institute of Medical Sciences Hapur, Panchsheel nagar, U.P., India
2 Department of Biochemistry, University College of Medical Sciences Delhi, India
3 Division of Pharmacology and Toxicology, Defence Research & Development Establishment, Gwalior, India
     

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Aims & Objective: In the present study our main objective was to develop a fluorescence sandwich ELISA assay for in vivo detection of ricin from plasma of mice after intoxication. Ricin (also called RCA-II or RCA60), one of the most potent toxins and documented bioweapons, is derived from castor beans of Ricinus communis.

Material & Method: The assay was performed in plasma obtained from ricin treated and control mice after 30 minutes of exposure. The assay utilizes an antiricin rat polyclonal antibody to adsorb ricin from plasma of ricin treated mice. The rabbit polyclonal antiricin antibody is then used to form a sandwich, and anti-rabbit FITC-conjugate allows development of fluorescence. The developed fluorescence was read at an excitation-emission wavelength of 485/528nm. The graph was drawn by plotting the ricin concentration against the fluorescence (rf) values.

Result: We were able to detect more than 19.5ng/ml of ricin in assay buffer, 28ng/ml in diluted plasma (1:20480) at the dose of 10mg/kg and 27ng/ml of diluted Plasma (1:80) at the dose of 5mg/ kg body weight. The coefficient of variation ranged from 22% for ricin in assay buffer, 9% for 5mg/ kg body weight dose plasma, 14% for 10mg/kg body weight dose plasma and 4% for non treated control plasma.

Conclusions: The detection of ricin from serum/plasma of individuals exposed to ricin will be quite difficult at lethal dose, estimated for humans to be 1 to 10μg per kg body weight following inhalation or injection. Hence, more efforts are needed for the detection of ricin at its lethal exposure dose.


Keywords

Ricin, In vivo Detection, Fluorescence Sandwich ELISA
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  • A Pilot Project to Develop in Vivo Detection of Ricin through Fluorescence Sandwich ELISA Technique

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Authors

Poonam Kachhawa
Department of Biochemistry Saraswathi Institute of Medical Sciences Hapur, Panchsheel nagar, U.P., India
Kamal Kachhawa
Department of Biochemistry, University College of Medical Sciences Delhi, India
Om Kumar
Division of Pharmacology and Toxicology, Defence Research & Development Establishment, Gwalior, India

Abstract


Aims & Objective: In the present study our main objective was to develop a fluorescence sandwich ELISA assay for in vivo detection of ricin from plasma of mice after intoxication. Ricin (also called RCA-II or RCA60), one of the most potent toxins and documented bioweapons, is derived from castor beans of Ricinus communis.

Material & Method: The assay was performed in plasma obtained from ricin treated and control mice after 30 minutes of exposure. The assay utilizes an antiricin rat polyclonal antibody to adsorb ricin from plasma of ricin treated mice. The rabbit polyclonal antiricin antibody is then used to form a sandwich, and anti-rabbit FITC-conjugate allows development of fluorescence. The developed fluorescence was read at an excitation-emission wavelength of 485/528nm. The graph was drawn by plotting the ricin concentration against the fluorescence (rf) values.

Result: We were able to detect more than 19.5ng/ml of ricin in assay buffer, 28ng/ml in diluted plasma (1:20480) at the dose of 10mg/kg and 27ng/ml of diluted Plasma (1:80) at the dose of 5mg/ kg body weight. The coefficient of variation ranged from 22% for ricin in assay buffer, 9% for 5mg/ kg body weight dose plasma, 14% for 10mg/kg body weight dose plasma and 4% for non treated control plasma.

Conclusions: The detection of ricin from serum/plasma of individuals exposed to ricin will be quite difficult at lethal dose, estimated for humans to be 1 to 10μg per kg body weight following inhalation or injection. Hence, more efforts are needed for the detection of ricin at its lethal exposure dose.


Keywords


Ricin, In vivo Detection, Fluorescence Sandwich ELISA

References