Development of mass-detection affinity biosensor for determination of prion proteins using dendrimers as a support for immobilisation specific antibodies

Majka Šnejdárková 1Alexandra Poturnayová 1Tibor Hianik 2

1. Institute of Animal Biochemistry and Genetics SASc, Moyzesova 61, Ivanka pri Dunaji 90028, Slovakia (Slovak Rep.)
2. Comenius University, Department of Biophysics and Chemical Physics, Mlynská dolina F1, Bratislava 842 48, Slovakia (Slovak Rep.)


Prion diseases are a group of fatal neurodegenerative disorders that include  scrapie in sheep , bovine spongiform encephalopathy, Creutzfeldt-Jakob disease and Gerstmann-Sträussler-Scheineker  diseases in humans . It is suggested that the main reason of these diseases  is asssociated with the  conversion of the  cellular  prion proteins (PrPC) into their misfolded  pathogenic isoform PrPSc. The accumulation of PrPSc   in the central nervous system is the common feature of the prion diseases. So far there is no available effective ante mortem test of the presence of PrPSc. At the same time, possibility of early diagnosis of prion disease could increase effectivity of the therapy. We therefore focused our work on development biosensors for detection prion proteins. For this purpose we used mass-sensitive affinity sensor based on quartz crystal microbalance (QCM).

Because PrPSc is highly infectious compound, we have focused on PrPC detection. The methods of detection of PrPSC and PrPC is, however analogical. The differences consists in using different antibodies. The PrPC specific antibodies PRI308 that selective bind to 106-126 amino acid residues of recombinat PrPC (recPrP) have been immobilised on a surface formed by mixture of with covalently attached neutravidin and tightly adsorbed biotinylated protein A. The protein A due to its natural afinity towards the Fc region of antibodies was used for the oriented immobilization of antibodies. We have shown, that QCM biosensor allowing to detect recPrP with detection limit 4.4 nM, which is sufficient for detection PrPC in blood (physiological concentrations 20 nM). However further effort would be necessary for development more sensitive method that will allow to detect trace concentration of PrPSc. These experiments are in progress.Time dependent AFM images performed in solution during interaction of recPrP with the PRI308 suggest that the surface roughness changed during the immobilization process. The integrity of the layer was investigated by cyclic voltammetry at the presence of redox couple Fe(CN)64-/3-. These measurements indicate that the sensitive layer with the PRI 308 is practically impermeable for redox probe species. 

Acknowledgements. This work was financially supported by European Commission under 6 FP program NoE NeuroPrion (Contract No. FOOD-CT-2004-506579), by Agency for Promotion Research and Development under the contract No. APVV-0362-07 and by Slovak Grant Agency (Projects No. 2/7134/27, VVCE-0064-07  and 1/4016/07 to T.H).

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Presentation: Poster at SMCBS'2009 International Workshop, by Majka Šnejdárková
See On-line Journal of SMCBS'2009 International Workshop

Submitted: 2009-08-21 09:47
Revised:   2009-08-21 09:49
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