Short communicationAn ultrasensitive squamous cell carcinoma antigen biosensing platform utilizing double-antibody single-channel amplification strategy
Introduction
Squamous cell carcinoma antigen (SCCA) is a kind of tumor antigen TA-4 separated from a cervical squamous cell carcinoma (SCC) (Takeuchi et al., 2003). The morbidity of cervical SCC increases from 12% to more than 90% with elevated circulating levels of SCCA (Kato, 1992). It is regarded as a sensitive tumor marker in patients with cervical cancer. Until now, several techniques have been developed in its detection, such as common sandwich immunosensor (Wu et al., 2013), enzyme-linked immunosensor assay (Erickson et al., 2010), and chemiluminescence immunoassay (Li et al., 2014b). All of these methods employ mesoporous materials to amplify signals with complex processes. Thus, a new strategy for the marker detection is necessary in the clinical test and therapeutic evaluation.
Immunoassay is an effective method in tumor marker detection (Ren et al., 2014a, Ren et al., 2014b). But the amplification strategy is mainly based on using the mesoporous nanomaterials (Lei and Ju, 2012, Wu et al., 2012, Zhang et al., 2011a) to enlarge the capacity of biomolecules and enhance signals (Bi et al., 2013, Jeong et al., 2013, Qu et al., 2014).
In this research, the double-antibody single-channel immunosensor was fabricated to realize the marker detection, which employed human immunoglobulin antibody (anti-HIgG) instead of mesoporous nanomaterials acting as the supporting framework to amplify the loading quantity of SCCA antibody (anti-SCCA) for the first time. In this strategy, the immunosensor would be able to achieve excellent properties by the following reasons. First, the immuno-active materials own better recognition capability to ensure the sensor specific recognition (Kiefel et al., 1987). Second, biomolecules have prominent hydrophilcity to form homogeneous solution than nanomaterials (Cheng and Rossky, 1998). Third, the size of antibody or antigen is about 10–15 nm which is much smaller than that of 100 nm (the traditional size of nanomaterials applied in immunosensor), so the nanomaterials (large scale) may not obtain a satisfied result due to an inadequate adsorption. Several smaller sized nanomaterials have been applied in the immunosensor in recent years (Feng et al., 2012a). Thus, the double-antibody single-channel immunosensor is an excellent candidate method to detect tumor markers.
In this research, gold nanoparticles (Au NPs) decorated with mercapto-functionalized graphene sheets (Au@GS) were synthesized to increase the fixing quantity of anti-HIgG. Pd icosahedrons were first used in immunosensor to amplify the antigen detection quantity. In geometry, typically, only one touch point exists between a sphere and another shape under the limited condition. In the area of immunosensor, spherical materials (such as noble metal, mesoporous materials, quantum dots) have been applied in the sensor frequently (Feng et al., 2012b, Li et al., 2014a, Zhang et al., 2011b). However, the point-of-touching may exhibit lower binding force than the spot-of-touching. The Pd icosahedrons own twenty spots and every spot may contact the biomolecules more tightly than the point-of-touching of nanosphere.
Section snippets
Experimental
In this work, mercapto-functionalized graphene sheets were synthesized to capture more Au NPs. And the testing system was three-electrode configuration (ESI†). The glassy carbon electrode (GCE) was polished with a series of alumina powders (1, 0.3, 0.05 μm) and washed with water. Afterwards, the Au@GS modified electrode could adsorb more anti-HIgGs to protect and strengthen the framework and enlarge the capacity of HIgG. Once BSA blocked the non-specific binding sites (other binding sites other
Results and discussion
GS–SH is synthesized according to a reported method (Wang et al., 2014) with some modification for the first time in this strategy (ESI†). The Au NPs are connected on the GS–SH sufficiently. From Fig. 2A, it can be clearly seen that the GS is silk-like and the morphology is straticulate. Au@GS is shown in Fig. 2B. In the margin and middle of the GS, there exist some brilliant points (a, c) and relative tint points (b, d). These points are all Au NPs indicating the GS is straticulate and the Au
Conclusion
In this research, a double-antibody single-channel amplification strategy was adopted to develop a novel immunosensor for the detection of SCCA. Pd icosahedrons were used as the connecter of anti-SCCA and anti-HIgG, which might increase the captured quantity of SCCA to realize a lower detection. Au@GS was applied in the fabrication to enlarge the captured biomolecules. The prepared immunosensor exhibited excellent properties which would provide a potential application in clinical diagnose.
Acknowledgments
This study was supported by the National Natural Science Foundation of China (Nos. 21175057, 21375047, 21377046, 21405059), the Science and Technology Plan Project of Jinan (No. 201307010), The Science and Technology Development Plan of Shandong Province (No. 2014GSF120004). And Qin Wei thanks the Special Foundation for Taishan Scholar Professorship of Shandong Province and University of Jinan (No. ts20130937).
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