1. Shahid Beheshti university 2. Shahid Beheshti university
Abstract
Introduction
Hepatitis b is an infectious disease caused by the hepatitis b virus (hbv) that affects the liver. recent estimates suggested that hbv infection caused 686 000 deaths in 2013. as 25% of people who acquire hbv as children will develop primary liver cancer or cirrhosis as adults, chronic hepatitis b virus infection (chb) is a major public health issue worldwide.
hbsag is the first serological marker to appear in the blood and is the most important marker of hbv infection. hbeag appears in the serum during the high replicative phase of hbv infection, which makes it a serum marker of active viral replication. hbeag is a viral protein, which is soluble and closely related to non-secretory capsid antigen hepatitis b core antigen (hbcag). the different immunoreactivity between hbeag and hbcag is created by a different conformation of the b cell epitopes. detection of hbsag in serum samples is commonly carried out using an immunoassay such as an enzyme-linked immunosorbent assay (elisa), which is complex to perform, time-consuming, and unsatisfactory for testing sensitivity. therefore, new methods for highly sensitive detection of hbv infection are urgently needed.
aptamers are specific recognition molecules with high affinity and specificity toward their targets. biosensors that employ aptamers as biorecognition elements are known as aptasensors. in this study, we select an hbsag-specific aptamer and use it to develop a new electrochemichal aptasensor based on reduced graphen oxide-functionalized gold nanoparticles.
Methods
In this work, we have developed an aptasensor platform for ultrasensitive hbsag detection based on hbsag-specific aptamer and reduced graphene oxide/au nano particles (rgoaunps). after fabrication of aptasensor electrode, its electrochemical response was studied using cyclic voltammetry technique.
synthesis of rgoaunps: graphene oxide was synthesized from commercial graphite according to hummers method with a minor change. the product, graphite oxide, was exfoliated in deionized water in an ultrasonic bath to form graphene oxide (go) nanosheets. by adding the product to water-dispersed au nanoparticles and sonication for 24 h at 40 °c, rgo/aunps were synthesized. the final product was filtered using a buchner funnel and washed with deionized water three times then dried at 50 °c for 12 h.
electrochemical measurements: a conventional three-electrode cell with an ag/agcl reference electrode (argental, 3 m kcl) were used in order to carry out electrochemical experiments.
Results
Our results indicated that the new hbsag aptasensor is highly sensitive for electrochemical detection of hbsag. the detection limit of this hbsag-detecting aptasensor is as low as 3ng/ml, which is much lower than the limit of a typical elisa used in hospitals. furthermore, this aptasensor works well and is highly specific to hbv infection.
Conclusion
In this work, electrochemical sensing of hbsag was studied using ssdna aptamer. our results show that this aptasensor can detect hbsag molecules preciously. electrochemical data illustrated that nucleotides sequence of aptamer were selected successfully. using rgo/aunps in the structure of biosensor electrode enhanced the conductivity and surface area of the electrode.
