Detection and precipitation of hepatitis B core antigenusing a fusion bacteriophage

Siti Salwa Hasmoni, Khatijah Yusoff, and Wen Siang Tan*

J. Gen. Appl. Microbiol., 51, 125-131(2005)

(Received June 23, 2004; Accepted January 12, 2005)

91394025

甘博介

Introduction-Phage ELISA

The phage enzyme-linked immunosorbent assay (phage-ELISA) is a modified ELISA-based technique with the primary antibody replaced with the fusion phage.

M13 phage has been chosen in this experiment, use E. coli ER2738 to prepare

The Life Cycle of M13

F-pilus

Phage Display Technology

利用轉殖將一段短片段的 peptide gene 附於噬菌體中，進而在不影響其感染力的情況下產生所需要的 peptide 於 phage 表面蛋白上以供利用

Use anti-HBcAg monoclonal antibody to select which phage has the highest interaction

將目標 peptide表現於 phage 的表面蛋白上

Peptide-WSFFSNI

A short peptide which is selected from phage peptide library

Has high affinity interaction with HBcAg(Hepatitis B core antigen)

Phage bearing the sequence WSFFSNI has the potential to be further developed as an immunodiagnostic marker

Results-Detection of HBcAg with fusion phage via phage/mAb ELISA.

HBcAg + Fusion Phage HBcAg + mAb

1:5000=1.6g/ml

Results-Detection of HBcAg with dot blot assay.

Fusion phage Monoclonal antibody

Results-Detection of HBcAg in HBV positive serum samples using phage ELISA

Results-Detection of HBcAg in HBV positive serum samples with fusion phage via phage-dot blot assay

1x1012pfu/ml

Discussion HBcAg has been one of the most important mar

kers for HBV infections. Production of monoclonal or antibodies raised

in cell lines or animals are costly and time consuming, which usually takes months

the phage bearing the WSFFSNI sequence interacts tightly to HBcAg with a dissociation constant (KD

rel) less than 25 nM

Conclusion

Recombinant M13 phage which is cheaper in terms of production and can be rapidly propagated.

The purification of the filamentous phage involves lesser steps compared to that of monoclonal antibody.

The Advantage of Phage-ELISA

This type of phage-based diagnostic reagent can save time, cost and energy required to produce monoclonal antibodies.

Higher sensibility than general amount of mAB (monoclonal antibody)

THE END

The type of phage display system

ELISA

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