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Western Blot Analysis of Organophosphate Exposure.

PB97207112

Publication Date	1997
Personal Author	Jeffrey, A. M.; Grassman, J.
Page Count	66
Abstract	A novel Western blot assay was developed which could detect acetylcholinesterase (AChE) from human erythrocytes. Isoelectric focusing (IEF) was used to separate at least five major isozymes of AChE. Inactivation of the catalytic activity with organophosphate pesticides did not change the isoelectric point of the isozymes, and therefore separation of native from organophosphate inhibited AChE did not occur. The degree of organophosphate inhibition did not influence the recognition of AChE by the monoclonal murine HR anti-ChEs. Native and organophosphate inactivated AChEs were not resolved by Western blotting that used alternative buffering systems. Treatment with organophosphates, diisopropylphophorofluoridate (5914) and paraoxon (311455) produced aged AChE as evidence by resistance to reactivation with pralidoximine. The authors conclude that the charge difference between native and organophosphorus treated and aged intact AChEs is not sufficient for separation by IEF or electrophoresis. However, an internally controlled protein based assay would eliminate both interindividual and intraindividual variability, thus making it possible to assess AChE inhibition in the absence of preexposure values.
Keywords	Pesticides Blood cells Cholinesterase inhibitors Poisoning Insecticides Organo-phosphorus Enzymes Erythrocytes Isozymes Western blot assay Acetylcholinesterase
Source Agency	National Institute for Occupational Safety and Health
Corporate Authors	Columbia Univ., New York. School of Public Health.; National Inst. for Occupational Safety and Health, Cincinnati, OH.; National Inst. of Environmental Health Sciences, Research Triangle Park,
Supplemental Notes	Prepared in cooperation with National Inst. of Environmental Health Sciences, Research Triangle Park, NC. Sponsored by National Inst. for Occupational Safety and Health, Cincinnati, OH.
Document Type	Technical Report
NTIS Issue Number	199724

Western Blot Analysis of Organophosphate Exposure.

Western Blot Analysis of Organophosphate Exposure.
PB97207112

Publication Date	1997
Personal Author	Jeffrey, A. M.; Grassman, J.
Page Count	66
Abstract	A novel Western blot assay was developed which could detect acetylcholinesterase (AChE) from human erythrocytes. Isoelectric focusing (IEF) was used to separate at least five major isozymes of AChE. Inactivation of the catalytic activity with organophosphate pesticides did not change the isoelectric point of the isozymes, and therefore separation of native from organophosphate inhibited AChE did not occur. The degree of organophosphate inhibition did not influence the recognition of AChE by the monoclonal murine HR anti-ChEs. Native and organophosphate inactivated AChEs were not resolved by Western blotting that used alternative buffering systems. Treatment with organophosphates, diisopropylphophorofluoridate (5914) and paraoxon (311455) produced aged AChE as evidence by resistance to reactivation with pralidoximine. The authors conclude that the charge difference between native and organophosphorus treated and aged intact AChEs is not sufficient for separation by IEF or electrophoresis. However, an internally controlled protein based assay would eliminate both interindividual and intraindividual variability, thus making it possible to assess AChE inhibition in the absence of preexposure values.
Keywords	Pesticides Blood cells Cholinesterase inhibitors Poisoning Insecticides Organo-phosphorus Enzymes Erythrocytes Isozymes Western blot assay Acetylcholinesterase
Source Agency	National Institute for Occupational Safety and Health
Corporate Authors	Columbia Univ., New York. School of Public Health.; National Inst. for Occupational Safety and Health, Cincinnati, OH.; National Inst. of Environmental Health Sciences, Research Triangle Park,
Supplemental Notes	Prepared in cooperation with National Inst. of Environmental Health Sciences, Research Triangle Park, NC. Sponsored by National Inst. for Occupational Safety and Health, Cincinnati, OH.
Document Type	Technical Report
NTIS Issue Number	199724