Detection and precipitation of hepatitis B core antigen using a fusion bacteriophage.
The nucleocapsids of hepatitis B virus (HBV) are made of 180 or 240 subunits of core proteins or known as core antigens (HBcAg). A fusion bacteriophage bearing the WSFFSNI sequence that interacts tightly to HBcAg was employed as a diagnostic reagent for the detection of the antigen using the phage-enzyme-linked immunosorbent (phage-ELISA), dot blot and immunoprecipitation assays. The results from phage-ELISA and dot blot assay showed that as low as 10 ng of HBcAg can be detected optimally by 1.0x10(12) pfu/ml fusion M13 bacteriophage. The sensitivity of the dot blot assay corresponds with that of the phage-ELISA. HBcAg in HBV positive serum samples can also be detected using the fusion phage via the phage-ELISA and phage-dot blot assay. The phage cross-linked to cyanogen bromide (CNBr) activated agarose can also be used to precipitate HBcAg in bacterial lysate. The optimum amount of phage needed for cross-linking to 1 g of agarose is about 7.0x10(6) pfu/ml which could also precipitate purified and unpurified HBcAg in bacterial lysate. This study demonstrates the potential of fusion bacteriophage bearing the sequence WSFFSNI as a diagnostic reagent and a ligand for the detection and purification of HBcAg respectively.
Hasmoni
Siti Salwa
SS
Department of Microbiology, Faculty of Biotechnology and Biomolecular Sciences, University Putra Malaysia, UPM 43400, Serdang, Selangor, Malaysia.
Yusoff
Khatijah
K
Tan
Wen Siang
WS
D017104:Bacteriophage M13 / Q000737:chemistry*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D006509:Hepatitis B / Q000175:diagnosis*; D006512:Hepatitis B Core Antigens / Q000302:isolation & purification*; D006515:Hepatitis B virus / Q000302:isolation & purification*; D006801:Humans; D015151:Immunoblotting; D047468:Immunoprecipitation; D011993:Recombinant Fusion Proteins / Q000737:chemistry*
D006512:Hepatitis B Core Antigens; D011993:Recombinant Fusion Proteins