Standardization of Histological Procedures for the Detection of Toxic Substances by Immunohistochemistry in Dipteran Larvae of Forensic Importance†
Carina M. Souza M.Sc.
Department of Animal Biology, IB, UNICAMP, Distrito de Barão Geraldo, Campinas, São Paulo, CEP 13083-970 Brazil
Search for more papers by this authorCarolina G. P. Lima M.Sc.
Department of Animal Biology, IB, UNICAMP, Distrito de Barão Geraldo, Campinas, São Paulo, CEP 13083-970 Brazil
Search for more papers by this authorMarcos J. Alves-Jr B.Sc.
Department of Animal Biology, IB, UNICAMP, Distrito de Barão Geraldo, Campinas, São Paulo, CEP 13083-970 Brazil
Search for more papers by this authorWagner W. Arrais-Silva Ph.D.
Campus of Araguaia, UFMT, Rodovia MT-100, Km 3,5, Pontal do Araguaia, Mato Grosso, CEP 78698-000 Brazil
Search for more papers by this authorSelma Giorgio Ph.D.
Department of Animal Biology, IB, UNICAMP, Distrito de Barão Geraldo, Campinas, São Paulo, CEP 13083-970 Brazil
Search for more papers by this authorArício X. Linhares Ph.D.
Department of Animal Biology, IB, UNICAMP, Distrito de Barão Geraldo, Campinas, São Paulo, CEP 13083-970 Brazil
Search for more papers by this authorCorresponding Author
Patricia J. Thyssen Ph.D.
Department of Animal Biology, IB, UNICAMP, Distrito de Barão Geraldo, Campinas, São Paulo, CEP 13083-970 Brazil
Additional information and reprint request:
Patricia J. Thyssen, Ph.D.
Department of Animal Biology
IB, UNICAMP
Distrito de Barão Geraldo
Campinas, São Paulo
CEP 13083-970
Brazil
E-mail: [email protected]
Search for more papers by this authorCarina M. Souza M.Sc.
Department of Animal Biology, IB, UNICAMP, Distrito de Barão Geraldo, Campinas, São Paulo, CEP 13083-970 Brazil
Search for more papers by this authorCarolina G. P. Lima M.Sc.
Department of Animal Biology, IB, UNICAMP, Distrito de Barão Geraldo, Campinas, São Paulo, CEP 13083-970 Brazil
Search for more papers by this authorMarcos J. Alves-Jr B.Sc.
Department of Animal Biology, IB, UNICAMP, Distrito de Barão Geraldo, Campinas, São Paulo, CEP 13083-970 Brazil
Search for more papers by this authorWagner W. Arrais-Silva Ph.D.
Campus of Araguaia, UFMT, Rodovia MT-100, Km 3,5, Pontal do Araguaia, Mato Grosso, CEP 78698-000 Brazil
Search for more papers by this authorSelma Giorgio Ph.D.
Department of Animal Biology, IB, UNICAMP, Distrito de Barão Geraldo, Campinas, São Paulo, CEP 13083-970 Brazil
Search for more papers by this authorArício X. Linhares Ph.D.
Department of Animal Biology, IB, UNICAMP, Distrito de Barão Geraldo, Campinas, São Paulo, CEP 13083-970 Brazil
Search for more papers by this authorCorresponding Author
Patricia J. Thyssen Ph.D.
Department of Animal Biology, IB, UNICAMP, Distrito de Barão Geraldo, Campinas, São Paulo, CEP 13083-970 Brazil
Additional information and reprint request:
Patricia J. Thyssen, Ph.D.
Department of Animal Biology
IB, UNICAMP
Distrito de Barão Geraldo
Campinas, São Paulo
CEP 13083-970
Brazil
E-mail: [email protected]
Search for more papers by this authorAbstract
Immunohistochemistry (IHC) technique is an alternative toxicological analysis to detect drugs in insects of forensic importance, but it requires thorough histological procedures. In this study, we tested different fixatives—phosphate-buffered paraformaldehyde 4% (PP), Carnoy's fluid (CF), Kahle's solution (KS), ethanol in different concentrations, and ethanol associated to PP and CF, time of fixation and histological processes for dipteran larvae's tissue, aiming to develop a sample preparation protocol for IHC application. A suitable fixation was achieved using PP for 12 and 24 h, CF for 3 h, 70% ethanol for 19 days, and 70% ethanol/CF for 2 h/3 h. Postfixation using negative pressure, two immersions in xylene for 30 min each, and one in xylene plus paraffin for 45 min increased tissue preservation. An immunohistochemical test for cocaine detection was performed using monoclonal benzoylecgonine antibody from mouse, peroxidase-conjugated anti-mouse IgG and visualized by 3,3′-diaminobenzidine method showed these histological procedures didn't compromise antigenicity.
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