Antibody detection employing sol-gel immobilized parasites.
Immunofluorescence assay (IFA) and immunoperoxidase assay (IPA) are useful diagnostic techniques for specific antibody detection for different diseases. Both involve several alternatives for immobilization of cells, such as solvent or heat fixation. Non-covalent immobilization implies rigorous storage conditions at -20 degrees C to preserve the slides, and usually numerous cells are detached during the washing steps, which can lead to inconsistencies in the results. Sol-gel chemistry is usually used for coating different materials because of the mild conditions of the polymerization reaction and the ability to introduce functional groups to a wide variety of surfaces. We have developed a novel procedure for the attachment of Trypanosoma cruzi epimastigotes and Leishmania guyanensis promastigotes to a silicon oxide polymer covered glass surface. The film was prepared using standard microscope slides with tetraethoxysilane and 3-aminopropyl triethoxysilane as polymeric precursors. When acetone was used as the major coating solvent, the IFA showed the fluorescence of the attached parasites without matrix background interference. Similar results were observed when the IPA was evaluated. The sensitivity and specificity of the sol-gel immobilized parasite slides were comparable with the heat fixation technique. The performance of the coated slides was maintained for at least 2 months at 4 degrees C storage temperature. This immobilization method does not affect the molecular epitopes of the attached cells. Thus, homogeneous, ready to use, long lasting coated slides were obtained, which are appropriate for field conditions.
Copello
Guillermo J
GJ
Cátedra de Química Analítica Instrumental, Facultad de Farmacia y Bioquímica, Universidad de Buenos Aires, Buenos Aires, Argentina.
De Marzi
Mauricio C
MC
Desimone
Martín F
MF
Malchiodi
Emilio L
EL
Díaz
Luis E
LE
D000096:Acetone / Q000737:chemistry; D000818:Animals; D000913:Antibodies, Protozoan / Q000097:blood*; D000431:Ethanol / Q000737:chemistry; D005455:Fluorescent Antibody Technique; D005782:Gels; D006801:Humans; D007124:Immunoenzyme Techniques; D018334:Leishmania guyanensis / Q000276:immunology*; D051379:Mice; D008807:Mice, Inbred BALB C; D011437:Propylamines; D015203:Reproducibility of Results; D012821:Silanes / Q000737:chemistry*; D012997:Solvents / Q000737:chemistry; D017550:Spectroscopy, Fourier Transform Infrared; D014349:Trypanosoma cruzi / Q000276:immunology*
D000913:Antibodies, Protozoan; D005782:Gels; D011437:Propylamines; D012821:Silanes; D012997:Solvents; D000096:Acetone; D000431:Ethanol; C040733:tetraethoxysilane; C477625:amino-propyl-triethoxysilane