title: Electrochemical immunosensor for detection of antibodies against influenza A virus H5N1 in hen serum.
abstract: This paper describes the development of an immunosensor for detection of anti-hemagglutinin antibodies. Its preparation consists of successive modification steps of glassy carbon electrodes: (i) creation of COOH groups, (ii) covalent immobilization of protein A with EDC/NHS coupling reaction, (iii) covering with anti-His IgG monoclonal antibody, (iv) immobilization of the recombinant His-tagged hemagglutinin (His6-H5 HA), (v) filling free space with BSA. The interactions between two variants of recombinant HA (short and long) from highly pathogenic avian influenza virus H5N1 and the anti-H5 HA monoclonal antibody (Mab 6-9-1) have been explored with electrochemical impedance spectroscopy (EIS). The impedimetric immunosensor displayed a very good detection limit (LOD) of 2.1 pg/mL, the quantification limit (LOQ) of 6.3 pg/mL and a dynamic range from 4 pg/mL to 20 pg/mL. In addition, this analytical device was applied for detection of antibodies against His6-H5 HA in serum of vaccinated hen using serial 10-fold dilutions of serum. The immunosensor proposed was able to detect antibody in hen serum diluted up to 7 × 10(7)-fold. The sensitivity of immunosensor was about four orders of magnitude much better than ELISA.
authors.affiliation: Institute of Animal Reproduction and Food Research of Polish Academy of Sciences, Tuwima 10, 10-748 Olsztyn, Poland.
authors.forename: Urszula
authors.initials: U
authors.lastname: Jarocka
authors.affiliation: Institute of Biochemistry and Biophysics, Polish Academy of Sciences, Pawińskiego 5A, 02-106 Warsaw, Poland.
authors.forename: Róża
authors.initials: R
authors.lastname: Sawicka
authors.affiliation: Institute of Biochemistry and Biophysics, Polish Academy of Sciences, Pawińskiego 5A, 02-106 Warsaw, Poland.
authors.forename: Anna
authors.initials: A
authors.lastname: Góra-Sochacka
authors.affiliation: Institute of Biochemistry and Biophysics, Polish Academy of Sciences, Pawińskiego 5A, 02-106 Warsaw, Poland.
authors.forename: Agnieszka
authors.initials: A
authors.lastname: Sirko
authors.affiliation: Institute of Biochemistry and Biophysics, Polish Academy of Sciences, Pawińskiego 5A, 02-106 Warsaw, Poland.
authors.forename: Włodzimierz
authors.initials: W
authors.lastname: Zagórski-Ostoja
authors.affiliation: Institute of Animal Reproduction and Food Research of Polish Academy of Sciences, Tuwima 10, 10-748 Olsztyn, Poland.
authors.forename: Jerzy
authors.initials: J
authors.lastname: Radecki
authors.affiliation: Institute of Animal Reproduction and Food Research of Polish Academy of Sciences, Tuwima 10, 10-748 Olsztyn, Poland. Electronic address: h.radecka@pan.olsztyn.pl.
authors.forename: Hanna
authors.initials: H
authors.lastname: Radecka
mesh_terms: D000818:Animals; D000914:Antibodies, Viral / Q000097:blood*; D015374:Biosensing Techniques / Q000295:instrumentation*; D002645:Chickens / Q000097:blood* / Q000276:immunology*; D003217:Conductometry / Q000295:instrumentation*; D004867:Equipment Design; D019544:Equipment Failure Analysis; D005260:Female; D019267:Hemagglutinin Glycoproteins, Influenza Virus / Q000276:immunology; D007118:Immunoassay / Q000295:instrumentation*; D053124:Influenza A Virus, H5N1 Subtype / Q000276:immunology*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
chemical_list: D000914:Antibodies, Viral; D019267:Hemagglutinin Glycoproteins, Influenza Virus
keywords: Avian influenza virus; Detection of antibody; Electrochemical impedance spectroscopy; Glassy carbon electrodes; Immunosensor