Nanotechnol Rev 2016; 5(4): 393–416 Review Deepshikha Saini* Synthesis and functionalization of graphene and application in electrochemical biosensing DOI 10.1515/ntrev-2015-0059 as “miracle material” for its outstanding characteris- Received October 13, 2015; accepted November 22, 2015; previously tics. Graphene is also the fundamental building block of published online February 4, 2016 carbon materials such as fullerenes, carbon nanotubes, and graphite [1, 2]. Its unique configuration and struc- Abstract: Graphene is a two-dimensional material with ture determine a number of fascinating properties such amazing characteristics, which grant it the title “wonder as a high planar surface area (2630 m2 g-1) [3], superior material”. It has grabbed appreciable attention due to its mechanical strength with a Young’s modulus of 1100 GPa exceptional electrical, optical, thermal, and mechani- [4], unparalleled thermal conductivity (5000 W m-1 K-1) [5], cal properties. Because of these interesting properties, remarkable high carrier mobility (10,000 cm2 V-1 s -1) [6], graphene has found its way into a wide variety of bio- high opacity (~97.7%), and the ability to quench fluores- sensing applications. It has been used as a transducer in cence [7]. Graphene is currently, without any doubt, the electrochemical biosensors, bio-field-effect transistors, most intensively studied material for a wide range of impedance biosensors, electrochemiluminescence, and applications that include electronics, energy, and sensing fluorescence biosensors. Functionalization of graphene outlets [8]. has further opened up novel fundamental and applied However, tuning physicochemical properties frontiers. The present article reviews recent works deal- becomes necessary [9] in many graphene applications ing with synthesis, functionalization of graphene, and because of intrinsic graphene properties like zero band its applications related to biosensors. Various synthesis gap, high sheet resistance, easy aggregation, and poor strategies, mechanism and process parameters, and types solubility, which are the big obstacles to the various of functionalization are discussed in view of biosensor applications [10, 11]. Functionalization is one of the effi- development. Some potential areas for biosensor-related cient ways to tailor the electrical properties of graphene. applications of functionalized graphene are highlighted, Furthermore, after modification, the as-synthesized including catalytic biosensors and bio affinity biosensors. hybrid materials could not only overcome the disadvan- Wherever applicable, the limitations of the present knowl- tages of intrinsic graphene but also be inculcated with edgebase and possible research directions have also been new desirable properties. Graphene is thought to become discussed. especially widespread in biosensors and diagnostics. The Keywords: electrochemical biosensors; functionalization; large surface area of graphene can enhance the surface graphene; synthesis. loading of desired biomolecules, and excellent conduc- tivity and small band gap can be beneficial for conduct- ing electrons between biomolecules and the electrode 1 Introduction surface. Graphene also has significant potential for ena- bling the development of electrochemical biosensors Graphene is composed of single-atom thick sheets of sp2 based on direct electron transfer between the enzyme bonded carbon atoms that are arranged in a perfect two- and the electrode surface. Moreover, other applications dimensional (2D) honeycomb lattice. It is often dubbed including, but not limited to, synthesizing nanoelectron- ics [12], high-frequency electronics [13], energy storage and conversion devices [14], field-emission displays [15], and transparent conductors [16] are also proofs of their *Corresponding author: Deepshikha Saini, Beijing National Laboratory for Molecular Sciences, Laboratory of Molecular versatility. Nanostructure and Nanotechnology, Institute of Chemistry, In this comprehensive review, recent research efforts Beijing 100190, P. R. China, e-mail: [email protected] to the synthesis, functionalization of graphene, and its 394 D. Saini: Functionalized graphene-based biosensors application in the development of next-generation bio- Mechanical exfoliation sensors are addressed. This covers the latest develop- (research, prototyping) ments and importantly, offer insights on the underlying CVD detection mechanisms and on the unique advantages of (coating, bio, transparent) conductive layers, graphene in comparison with other materials. I hope that electronics, this article would inspire broader interests across various photonics) disciplines and stimulate more exciting developments in this still young, yet very promising, field of research. Quality SiC (electronics, Molecular RF transistors) assembly 2 Synthesis of graphene (nanoelectronics) As the first step to realize the applications, graphene must Liquid-phase exfoliation (coating, composites, be synthesized on a large scale with controlled size and inks, energy storage, morphology. Controllable and scalable production of gra- bio, transparent conductive layers) phene is of great importance for its applications in dif- Price (for mass production) ferent fields. Graphene has been synthesized in various Figure 1: Methods for the mass production of graphene. There ways and on different substrates. In the following, we are several choices depending on the particular application, each summarize the synthesis methods to obtain high-quality with differences in terms of size, quality (e.g. presence of defects graphene. The methods for graphene preparation can be and impurities), and price. Reprinted with permission from Ref. [8] divided into several groups: (Copyright 2012 Nature). (i) Mechanical exfoliation of graphene layers from highly oriented pyrolytic graphite [17–19] (i) Capability of automation and miniaturization (ii) Chemical vapor deposition with hydrocarbon decom- (ii) High electron transfer rate position on a metal [20–24] (iii) Biocompatibility (iii) Thermal decomposition of SiC (epitaxial growth) (iv) Easier operation [25–29] (v) Extraordinary carrier mobility and capacitance (iv) Electrical arc discharge method [30–33] (vi) Excellent transparency (v) Organic synthesis [34–37] (vii) Exceptionally large surface-to-volume ratio (vi) Chemical method using dispersion of graphite. (viii) Single atom thickness [38, 39] (ix) Lower cost (x) Robustness and flexibility The mainstream methods of graphene synthesis have (xi) Excellent conductivity been summarized in Figure 1. Table 1 summarizes the relative advantages and dis- These properties have also led to significant interest in advantages of the above synthesis methods in terms of developing graphene-based electrochemical biosensors the feasibility to scale-up the process for mass production, [48]. Graphene has been employed as electrode material materials and production costs, and the presence of defects. in various electrochemical biosensors for the detection of a range of analytes like glucose, glutamate, choles- terol, hemoglobin, and more [49]. Interestingly, the 2.1 Significance of graphene for electro- large surface area of and excellent conductivity leads chemical biosensing to the superior electrochemical biosensing performance of graphene over carbon nanotubes in terms of sensi- Graphene and sensors are a natural combination, as gra- tivity, signal-to-noise ratio, electron transfer kinetics, phene’s large surface-to-volume ratio, unique optical and stability [50, 51]. Additionally, graphene’s proper- properties, excellent electrical conductivity, high carrier ties can be tuned by amendable synthetic conditions, mobility and density, high thermal conductivity, and dimensions, number of layers, and doping components many other attributes can be greatly beneficial for sensor [52]. Therefore, graphene-based materials are playing, functions. The superior advantages offered by graphene and will continually play, a significant role in sensing for electrochemical biosensing are as follows applications. Table 1: Comparison of different graphene synthesis methods. Sr. No. Synthesis methods Size Advantages Disadvantages Applications References 1. Mechanical Flakes (5 to 100 μm) Simplicity, high quality, less Delicate, low yields, uneven Basic research purpose [18, 19] cleavage defects films, not scalable 2. CVD (on Ni, Cu Co) Thin films ≤ 75 cm Scalable, high quality, uniform, High process temperature Touch screens, smart [22–24] high compatibility ( > 1000°C), high cost, windows, flexible LCDs, complex transfer process OLEDS, and solar cells 3. Epitaxial growth Thin films ( > 50 μm) Large-scale production, high High process temperature Graphene electronics [28, 29] on SiC quality, no defects (1500°C), high cost, low yields, discontinuous 4. Chemical reduction Nanoflakes/powder High scalability, high yields, High defect density, low Conductive inks and [40–42] of graphite oxide (nm to a few μm) low cost, easily processable purity paints, polymer fillers, biosensors graphene-based Saini: Functionalized D. super capacitors, sensors 5. Liquid phase Nanosheets (nm to Simplicity, high scalability, low Moderate quality, impure, Transparent electrodes, [43, 44] exfoliation a few μm) cost low yield sensors, polymer fillers 6. Carbon nanotubes Nanoribbons (few Low-cost, high quality, high Time-consuming, moderate FET interconnects,NEMs [45, 46] unzipping microns) yield scalability composite s 7. Arc discharge of Nanosheets (100s High crystallinity,