title: Detection of Rubisco and mycotoxins as potential contaminants of a plantibody against the hepatitis B surface antigen purified from tobacco.
abstract: Antibodies have been one of the proteins widely expressed in tobacco plants for pharmaceutical purposes, which demand contaminant free preparations. Rubisco constitutes 40-60% of tobacco leaf soluble proteins; therefore it is the major potential protein contaminant of plantibodies, while mycotoxins are toxic compounds that could be introduced during the biomass production and post-harvest stages with important consequences to human health. The objective of this paper was to investigate whether Rubisco and mycotoxins are present in Plantibody HB-01 preparations used in the immunopurification of the hepatitis B surface antigen. Rubisco was purified from Nicotiana tabacum yielding 154 microg of protein per gram of leaves and purity over 95%. Among mouse monoclonal antibodies generated against this enzyme, the CBSS.Rub-2 was selected for its immunodetection. It recognizes a conserved sequential epitope of Rubisco large subunit with an affinity constant of 0.13 x 10(8)M(-1). Rubisco quantification limit was 1 microg spreading to the measurement of this contaminant less than 4% of plantibodies samples. Additionally, according to a Reverse Phase-HPLC used to measure the level of adventitiously introduced contaminants, it can be concluded that aflatoxins B1, B2, G1 and G2 were undetected in the purified Plantibody HB-01 samples.
authors.affiliation: Tobacco Research Institute, Tumbadero Road 8.5 km, San Antonio de los Baños, Havana, La Havana, Cuba. dgeadalop@yahoo.com.mx
authors.forename: Déborah
authors.initials: D
authors.lastname: Geada
authors.forename: Rodolfo
authors.initials: R
authors.lastname: Valdés
authors.forename: Arturo
authors.initials: A
authors.lastname: Escobar
authors.forename: Dulce M
authors.initials: DM
authors.lastname: Ares
authors.forename: Edel
authors.initials: E
authors.lastname: Torres
authors.forename: Reinaldo
authors.initials: R
authors.lastname: Blanco
authors.forename: Williams
authors.initials: W
authors.lastname: Ferro
authors.forename: Dayamí
authors.initials: D
authors.lastname: Dorta
authors.forename: Marcos
authors.initials: M
authors.lastname: González
authors.forename: María R
authors.initials: MR
authors.lastname: Alemán
authors.forename: Sigifredo
authors.initials: S
authors.lastname: Padilla
authors.forename: Leonardo
authors.initials: L
authors.lastname: Gómez
authors.forename: Norma
authors.initials: N
authors.lastname: Del Castillo
authors.forename: Otto
authors.initials: O
authors.lastname: Mendoza
authors.forename: Dioslaida
authors.initials: D
authors.lastname: Urquiza
authors.forename: Yordanka
authors.initials: Y
authors.lastname: Soria
authors.forename: José
authors.initials: J
authors.lastname: Brito
authors.forename: Alberto
authors.initials: A
authors.lastname: Leyva
authors.forename: Carlos
authors.initials: C
authors.lastname: Borroto
authors.forename: Jorge V
authors.initials: JV
authors.lastname: Gavilondo
mesh_terms: D000348:Aflatoxins / Q000032:analysis* / Q000737:chemistry; D000818:Animals; D000911:Antibodies, Monoclonal / Q000235:genetics / Q000302:isolation & purification; D015153:Blotting, Western; D002851:Chromatography, High Pressure Liquid / Q000379:methods / Q000706:statistics & numerical data; D004340:Drug Contamination; D004797:Enzyme-Linked Immunosorbent Assay; D006510:Hepatitis B Antibodies / Q000235:genetics / Q000302:isolation & purification*; D006514:Hepatitis B Surface Antigens / Q000276:immunology; D006801:Humans; D051379:Mice; D046549:Plantibodies / Q000235:genetics / Q000302:isolation & purification*; D030821:Plants, Genetically Modified; D012273:Ribulose-Bisphosphate Carboxylase / Q000032:analysis*; D014026:Nicotiana / Q000235:genetics / Q000276:immunology
chemical_list: D000348:Aflatoxins; D000911:Antibodies, Monoclonal; D006510:Hepatitis B Antibodies; D006514:Hepatitis B Surface Antigens; D046549:Plantibodies; D012273:Ribulose-Bisphosphate Carboxylase