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, 2007. Original Russian Text c G.K. Chudinova, I.A. Nagovitsyn, M.A. Kononov, I.A. Maslyanitsyn, V.D. Shigorin, V.V. Savranskii, 2007, published in Kratkie Soobshcheniya po Fizike, 2007, Vol. 34, No. 12, pp. 43–51.
Atomic-Force Microscopy Studies of the Complexes of DNA and Single-Wall Carbon Nanotubes
G. K. Chudinova, I. A. Nagovitsyn, M. A. Kononov, I. A. Maslyanitsyn, V. D. Shigorin, and V. V. Savranskii
Prokhorov General Physics Institute, Russian Academy of Sciences, ul. Vavilova 38, Moscow, 119991 Russia
Received November 19, 2007
Abstract—The interaction of DNA samples extracted from tobacco leaves of two types with carbon nanotubes (CNTs) was studied. Atomic-force microscopy images of DNA–CNT complex ﬁlms were obtained. The possibility of forming ordered self-assembling structures in a DNA–CNT ﬁlm was shown. DOI: 10.3103/S1068335607120068
Recently, carbon nanotubes attract close attention, in particular, as biocompatible materials with biological and medical applications. Carbon nanotube-based bionanocomposites can be applied to (i) biosensor production; (ii) synthesis of molecular structures for vaccine and drug transport to a certain human organ; (iii) the use of nanotubes as a cell growth matrix; (iv) the development of bioelectronic devices which can be implanted into the human organism [1–5]. The use of nanotubes in the development of a sensitive element of biosensors leads to appreciable advantages, speciﬁcally, a probably sharp sensitivity enhancement, a decrease in the sensitive element size due to a developed surface, and an increase in the measurement rate and reproducibility [6–13]. Recently, atomic-force microscopy (AFM) is successfully used as a detecting element of a sensor. This method has giant potential for both quantitative and qualitative determination of biomaterials. AFM is a very accurate instrument for characterizing the image of complex biomolecular systems. Carbon nanotubes (CNTs) represent an ideal material for these purposes. Recently, AFM in the dynamic mode is used for chemical and molecular-biological analyses of DNA and not only DNA [13–27]. This study is the ﬁrst among those of the planned series with the overall aim to perform a systematic study on the choice of optimum conditions of nanotube-based bionanocomposite fabrication for the sensitive element of the biosensor. Experimental. Single-wall raw nanotubes (NTs) synthesized by the technique  were used; the nanotube content in the raw product was 20%. DNA samples (DNA1 and DNA2) were extracted from leaves of two tobacco sorts of genuine Nicotiana Tabacum L. by the technique . The initial DNA concentrations were 1.5 · 10−6 M (DNA1) and 3.3 · 10−6 M (DNA2). It is known from the literature that the extraction of nanotubes themselves from a reaction mixture is performed using solutions of surfactants such as triton Х-100 with various concentrations [30, 31]. In this study, DNA molecules themselves were used as an active material for NT extraction from the initial product. DNA–NT complexes were obtained in water (pH 6.0) and in a 0.05-M carbonate- bicarbonate buﬀer (рН 9.2) containing sodium chloride with a concentration of 0.02 M. The experiment was carried out as follows: four weighed portions of raw NTs (2 mg) were poured with hot solutions (95◦ C) containing DNA1 or DNA2 in amounts of 200 µl in 6 ml of water or buﬀer. The prepared samples were exposed to ultrasound using an UZDN-F device for 90 min. The DNA–NT solutions were cooled during ultrasound processing using a mixture of ice, sodium chloride, and water (the initial temperature was –7◦ C). The DNA and NT temperature during this process was on average 35◦ C. The prepared suspension was centrifuged for 20 min at 2000 rpm using an OP-8UKhL4.4 centrifuge. The solution containing the DNA–NT complex was placed using a micropipet (with a maximum volume of 200 µl) onto polished
7 and 8.5 mm in size) preliminarily placed into Petri dishes. Figure 1 shows the absorption spectra of initial concentrated DNA1 and DNA2 solutions (a) and the absorption spectra of the obtained DNA complex with NTs after ultrasonic processing in solution (b). The ﬁlm structures of the DNA1 complexes with NT (Fig. obtained by evaporating an aqueous solution at рН 6. The determination of the structure of the BULLETIN OF THE LEBEDEV PHYSICS INSTITUTE Vol. 34 No. Results and discussion. Russia) in the force scanning mode using the P7− SPM program. The ﬁlm structure was studied using a Р4-SPM-MDT microscope (ZAO “NT-MDT” Company. 6). DNA–NT complex solutions. This suggests that the shift of the DNA absorption band maximum during the interaction with NTs is explained exactly by the formation of the DNA complex with NTs. We can see the amorphous structure of the initial sample. The structure of ﬁlms containing DNA2 complexes with NTs represent aggregates of ball (100–120 nm) and oval (150– 350 nm) structures (Fig. the size of a single domain (“branch”) is 300 nm and larger.05-M buﬀer solution). (a) Absorption spectra of initial concentrated solutions of DNA1 and DNA2 before mixing with nanotubes. respectively. 3 (before ultrasonic extraction). Plates were dried for a day in closed Petri dishes at room temperature. 2). The electron microscopy image of raw NTs (the initial product containing NTs) is shown in Fig. and ﬁlms on quartz plates were measured using a Shimadzu UV 250 spectrophotometer. the absorption spectrum maximum of the DNA complex with NTs redshifts by 15–20 nm with respect to the DNA absorption maximum both in solution (Fig. For DNA1 (Fig.ATOMIC-FORCE MICROSCOPY 367 Fig. which does not allow us to suggest that there is only one single-wall NT in the DNA1 complex with CNT.0 after ultrasonic processing. rather than by a change in the environment during the ﬁlm formation. 5) remind a spruce branch. in the presence of 0. 1) and on the quartz substrate (Fig. The cord width is about 100 nm. 4). 1. braided with the DNA1 molecule (the schematic diagram of the complex is shown in [35. which are shown in Figs. obtained by evaporating a buﬀer solution (рН 9. 5 and 6) in comparison with the singlecomponent DNA1 ﬁlm structure (Fig. Because of the small sizes of NTs themselves [32–34]. As can be seen. Figure 4 shows the AFM image of ﬁlm grown by evaporation of DNA1 solution without NT addition on a quartz plate (control experiment).02-M sodium chloride). 7). We can see an essential diﬀerence in both size and shape of the structures formed in the ﬁlms (Figs. AFM imaging of the surface consisting of NTs has some diﬃculties. Quite diﬀerent images are characteristic of DNA1 and DNA2 complexes with NTs. quartz substrates (35×10×1. (b) absorption spectra of DNA1 and DNA2 with nanotubes after ultrasonic processing (200 µl of initial concentrated solution of DNA1 or DNA2 + 2 mg of raw nanotubes in 6 ml of 0. The absorption spectra of initial DNA solutions. Figure 2 shows the absorption spectra of initial DNA samples and DNA complexes with NTs on quartz substrates after ultrasonic processing and drying in air. 36]). highly ordered cord-like structures uniformly distributed over the surface are observed. Figures 5 and 6 show the images of NT ﬁlms with DNA1 and DNA2.2. 12 2007 . It is easily seen that the surface consists of densely packed ball structures which diameter varies from 30 to 60 nm.
as this is often observed for Langmuir ﬁlms. This can be caused. second. 2. obtained complex requires further study. AFM image of the DNA1 ﬁlm (without nanotubes). ﬁrst. the so-called defects. the ﬁlm structure represents domains with voids between them. 4. Fig. We note that atomic-force scanning of the quartz plate shows inhomogeneity of the obtained ﬁlm in all cases. Fig. Absorption spectra of two DNA samples without nanotubes (1 and 3) and DNA complexes with nanotubes (2 and 4) on the quartz substrate. Electron microscopy image of raw nanotubes. 5. 3. Fig. AFM image of the DNA1 ﬁlm with nanotubes. by that it is impossible to prepare a uniform ﬁlm by solution evaporation. 12 2007 .368 CHUDINOVA et al. The product contains 20% of nanotubes. A decrease in BULLETIN OF THE LEBEDEV PHYSICS INSTITUTE Vol. 34 No. Fig. obtained by evaporating an aqueous solution.
Similar highly ordered systems can be further used in the development of sensitive elements of sensor systems. BULLETIN OF THE LEBEDEV PHYSICS INSTITUTE Vol. AFM image of the DNA2 ﬁlm with nanotubes. 6. by the example of DNA samples extracted from leaves of tobacco of two diﬀerent sorts. defects of such a ﬁlm by the approach of domains results in ﬁlm destabilization (destruction). ﬁlamentary structures are formed (Fig. obtained by evaporating a buﬀer solution (рН = 9.2). Magniﬁcation of the image (Fig. However. such a structure of domain and voids alternation is thermodynamically favorable [37–40]. AFM image of the DNA1 ﬁlm with nanotubes. It was shown that selfassembled ordered domains consisting of DNA–CNT complexes are formed on the quartz plate surface.ATOMIC-FORCE MICROSCOPY 369 Fig. We note that ball  and ﬁlament structures are described in the literature for various DNA types . self-assembling of DNA1 complexes with CNTs during solution evaporation is the fact. Fig. 12 2007 . 8a). probably. obtained by evaporating an aqueous solution. 8b) shows that ﬁlamentary structures are composed of ball domains 100 nm in size and larger. 34 No. 7. it was shown that the diﬀerence in the nucleotide composition of DNA has an eﬀect on the shape of aggregates of DNA complexes with CNTs formed on the quartz plate surface.2). In the case of DNA2 (рН=9. We assume that aggregation of DNA2 complexes and nanotubes occurs in observed domains (aggregation of nanotubes was demonstrated in  by AFM). Thus.
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