Co-immobilized poly(ethylene glycol)-block-polyamines promote sensitivity and restrict biofouling on gold sensor surface for detecting factor IX in human plasma

In order to detect an extremely low amount of human coagulation factor IX (FIX), poly(ethylene glycol) (PEG)/aptamer co-immobilized surface was constructed using original PEG–polyamine surface modification agents on surface plasmon resonance (SPR) sensor chip. Initially, a gold (Au) sensor chip of SPR was modified using poly(ethylene glycol)-b-poly[2-(N,N-dimethylamino)ethyl methacrylate] (PEG-b-PAMA) followed by treatment with SH-dT20 and was duplexed with anti-FIX aptamer extended using A24. Furthermore, the co-immobilization of pentaethylenehexamine-terminated poly(ethylene glycol) (N6-PEG) on the sensing surface completely quenched bio-fouling. On this dual tethered PEG-surface, we determined that the dissociation constant for FIX–aptamer interaction was 37 ± 10 pM, and the sensitivity of detection could reach up to 800 fM on using aptamer–FIX–antibody sandwich pattern detected by gold nanoparticle-conjugated anti-mouse antibody. We could detect FIX in the presence of abundant albumin. Furthermore, to mimic the actual detection of FIX in clinical samples, we demonstrated our experimental results with human blood plasma instead of FIX. Higher-sensitivity was attained because of dual polymers immobilized on Au surface, and this can emerge as a common strategy for any aptamer–protein interactions. The selective binding of aptamer in human blood plasma described here indicates the suitability of the present strategy for detection in clinically relevant samples.