Letter www.acsami.org
Fabrication of Carbohydrate-Conjugated Fingerprintlike Mesoporous Silica Net for the Targeted Capture of Bacteria Nanjing Hao,*,†,‡ Laifeng Li,‡ and Fangqiong Tang*,‡ †
Thayer School of Engineering, Dartmouth College, Hanover, New Hampshire 03755, United States Technical Institute of Physics and Chemistry, Chinese Academy of Sciences, Beijing 100190, P. R. China
‡
S Supporting Information *
ABSTRACT: Herein, a rapid, straightforward, reliable, and low-cost strategy for targeted capture and detection of bacteria using carbohydrate-conjugated mesoporous silica structure was developed. Fingerprint-like mesoporous silica net (FMSN) with well-defined three-dimensional architecture and ordered morphology was first facilely synthesized by the aid of tetrabutylammonium iodine (TBAI) as cotemplates with cetyltrimethylammonium bromide (CTAB). When conjugated with maltoheptaose as targeting moiety, FMSN showed efficient and selective capturing capability of Staphylococcus epidermidis. This new and unique platform for capturing S. epidermidis is fast (within 18 min), high efficiency (greater than 98.6% from 1 × 103 CFU/mL to 1 × 108 CFU/mL), specific (compared to M. smegmatis mc2 155), and reusable (6 cycles). KEYWORDS: carbohydrate, mesoporous silica, fingerprint, targeting, bacteria
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Herein, we report an one-pot strategy for a scalable synthesis of fingerprintlike mesoporous silica net (FMSN), which possessed well-defined three-dimensional structure and ordered morphology. This unique FMSN structure was formed by the addition of tetrabutylammonium iodine (TBAI) as a cotemplate together with cetyltrimethylammonium bromide (CTAB). When maltoheptaose (G7), which contains seven glucose units linked by α-1,4-glycosidic bonds, was conjugated on the FMSN scaffold, the resulting FMSN-G7 multivalent system showed strong and selective binding interactions toward Staphylococcus epidermidis (S. epidermidis) (Figure 1). Fingerprintlike mesoporous silica material was synthesized using CTAB and TBAI as cotemplates in the presence of aqueous ammonia as a catalyst and tetraethyl orthosilicate (TEOS) as a silica source (See experimental details in ESI). As shown in Figure 2, well-defined FMSN material was successfully obtained in a large scale. The macroscopic morphology of FMSN is regular square or nearly circle (Figure 2A). These FMSNs have an average thickness of ∼100 nm (98 ± 13 nm, Figure S1), and an average diameter of ∼20 μm (21 ± 10 μm). In each FMSN, there are generally 20−50 concentrically arranged fibers (Figure 2B, C). The width of each fiber is ∼100 nm and the gap distance between fibers is ∼150 nm (Figure 2D, E). In addition, each fiber has mesoscopic structure, and the mesoporous channels are vertical to the longitudinal plane of the fiber (Figure 2F). Nitrogen
acteria can lead to environmental contamination and even serious disease, which bring a high public health burden over time. Rapid, straightforward, and reliable capture and detection of pathogenic bacteria is a key step in preventing the spread of contamination and infection. To date, bacterial detection methods include immunoassays, culture, and nucleic acid amplification (polymerase chain reaction, PCR).1−4 However, these methods are often time-consuming, laborintensive, or technically challenging, which greatly limited their widespread use, especially in resource-limited regions. Thus, new approaches that are fast, simple, reliable, and low-cost to capture and detect bacteria are still in high demand. Carbohydrates are found ubiquitously in nature as building blocks of the structural framework of bacteria and cells, as sources of metabolic energy, and as key components of various intercellular recognition processes.5 Carbohydrate-conjugated glycomaterials from organic and inorganic sources have recently been developed for cell tracking and classification, pathogen recognition, and in vivo diagnosis and therapy.6−8 These functions rely on the multivalent presentation of carbohydrate ligands on the nanomaterials, which amplifies the binding affinity between an individual monosaccharide and the corresponding receptor by orders of magnitude. However, developing bacterial capture and detection system that based on glycomaterials in a facile and specific way has still been proven challenging.9 On the other hand, since the discovery of mesoporous silica materials, great efforts have been made to alter their morphologies for meeting the increasing needs of maximum applications efficacy, especially in theranostic fields.10−13 © XXXX American Chemical Society
Received: August 31, 2016 Accepted: November 4, 2016
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DOI: 10.1021/acsami.6b10989 ACS Appl. Mater. Interfaces XXXX, XXX, XXX−XXX
Letter
ACS Applied Materials & Interfaces
To explore the formation mechanism of this unique fingerprintlike net structure and the pore orientations, we carried out a series of experiments to reveal the particle evolution process. Without the addition of TBAI, typical spherical mesoporous silica nanomaterial was formed (Figure S3). When TBAI was replaced with tetramethylammonium iodine (TMAI) or tetraethylammonium iodine (TEAI), fiberlike mesoporous silica structure was also formed, but there was no well-defined net structure (Figures S4 and S5). These results indicate that TBAI not only helps to construct fingerprintlike net architecture but also regulates the pore channels arrangement. The amount of TEOS also plays a significant role (Table S1). Slightly increasing the amount of TEOS reduced the gap distance between fibers, but the fiber contours were still visible under microscope (Figure S6). The fibers disappeared with a continued increase in TEOS and particles started to display a flat and homogeneous structure (Figure S7), which is similar to previous plate-like structure but with unique inherent characteristics.10,15 These obvious structural differences further confirmed the important roles of TEOS and revealed the formation process of FMSN (Table S1). On the basis of these observations, we propose that the growth of the FMSN involves two stages. The first stage is the cooperative assembly between surfactant molecules and silica species to form nucleation sites; the second stage is the continuous growth of the nucleation sites to nanofibers and the resultant fingerprintlike mesoporous net structure. In the first stage, CTAB and TBAI as cotemplates form rod-shaped long micelles, which then spontaneously bend to loops by lowering the surface energy.16 Combined with the considerations of interfacial energy between the micelles and the electrical charge interactions of the micellar systems, square- or circle- shaped structural arrangement was created.17 Silica species from the hydrolysis of TEOS assist in the formation of a hexagonally packed array of pore channels oriented perpendicular to the fiber length.18,19 The second stage is related to the side growth, during which surfactant loops and silica species coassemble along the side of nanofibers and thus fibers grow thicker.20 TBAI in micelle loops could generate surface repulsive forces, which is responsible for producing of gap distance between two fibers.17,21 Finally, fingerprintlike net with perpendicular pore channels architecture was obtained. The unique three-dimensional architecture, ordered morphology and pore channels, and large specific surface area make FMSN a promising new material. To explore its potential applications, we next used FMSN to capture and detect bacteria. Staphylococcus epidermidis (S. epidermidis) is a spherical bacterium and an important opportunistic pathogen. S. epidermidis is the most frequent cause of nosocomial infections and represents the most common source of infections on indwelling medical devices.22 Developing an effective and specific strategy to capture and detect S. epidermidis is therefore urgently needed. To realize this, maltoheptaose (G7), one kind of maltodextrin, which is especially metabolized and transported by S. epidermidis,22−26 was conjugated on FMSN using previously developed photocoupling chemistry by treating FMSN with PFPA-silane followed by photolysis in the presence of G7 (See details in the Supporting Information).6,27 The conversion from FMSN to G7-conjugated FMSN (FMSN-G7) was monitored by FTIR (Figure S8). The conjugation density of G7 on FMSN-G7 was ∼4.7 wt %, determined by thermal gravimetric analysis (Figure S9).
Figure 1. Fabrication of FMSN-G7 platform for targeted capture of S. epidermidis.
Figure 2. (A, B) SEM and (C−F) TEM images of FMSN at different magnifications.
adsorption−desorption measurement showed that FMSN exhibited typical type IV isotherm with H3 hysteresis (Figure S2), which is expected for FMSN with ordered mesoporous cylindrical pores and the presence of macroporous structure.14 The pore size of FMSN was calculated to be 3.6 nm by the Barrett−Joyner−Halenda (BJH) method. The Brunauer− Emmett−Teller (BET) surface area and pore volume were measured to be 258.3 m2/g and 0.51 cm3/g, respectively. B
DOI: 10.1021/acsami.6b10989 ACS Appl. Mater. Interfaces XXXX, XXX, XXX−XXX
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ACS Applied Materials & Interfaces
fully detached from particle surface. Results showed that similar bacteria capture performance was achieved within 6 cycles of treatment (Figure S11), reflecting a good reusability of glycomaterials.6,28 To further investigate the interactions between bacteria and FMSN-G7, we used confocal fluorescence microscopy. As shown in Figure 4, many red fluorescent SYTO 61-stained S.
To demonstrate the selective capture ability of FMSN-G7 toward S. epidermidis, M. smegmatis mc2 155 (rodlike bacteria) were used as a control. To facilitate the visualization of the capture process, bacteria were stained with SYTO 61, a nucleic acid fluorescent dye. As shown in Figure 3A, after SYTO 61 dye
Figure 3. (A, B) Optical images before and after particles were treated with bacteria for 2 h, respectively. Bacteria were stained with SYTO 61 dye. (a) 1 × 108 CFU/mL S. epidermidis; (b) 1 × 108 CFU/mL M. smegmatis mc2 155; (c) 1 × 108 CFU/mL S. epidermidis with 0.05 mg/ mL FMSN; (d) 1 × 108 CFU/mL M. smegmatis mc2 155 with 0.05 mg/mL FMSN; (e) 1 × 108 CFU/mL S. epidermidis with 0.05 mg/mL FMSN-G7; (f) 1 × 108 CFU/mL M. smegmatis mc2 155 with 0.05 mg/ mL FMSN-G7. (C) Capturing kinetic profile of S. epidermidis (1 × 108 CFU/mL) treated with FMSN-G7 (0.05 mg/mL). (D) Capturing efficiency profile of different concentration of S. epidermidis treated with 0.05 mg/mL FMSN-G7. Results were obtained in triplicates.
Figure 4. Confocal microscopy images of (A) S. epidermidis and (B) M. smegmatis mc2 155 treated with FMSN-G7. Images from left to right panel are bright-field, red channel, and overlay, respectively. Bacteria were stained with SYTO 61 dye.
epidermidis bacteria were observed on FMSN-G7 surface after incubation for 30 min, indicating that S. epidermidis efficiently bound on FMSN-G7. In comparison, no obvious fluorescence was observed when M. smegmatis mc2 155 was used (Figure 4B). These results are in agreement with the observations from Figure 3, which further demonstrated that the capture of S. epidermidis by FMSN-G7 is specific, fast, and effective. In summary, we developed a rapid, straightforward, reliable, and low-cost strategy for targeted capture and detection of S. epidermidis using maltoheptaose-conjugated fingerprintlike net material. FMSN with unique three-dimensional architecture and ordered pore channels was synthesized simply by using CTAB and TBAI as cotemplates. FMSN-G7 as multivalent platform for capturing S. epidermidis is fast (within 18 min), high efficiency (greater than 98.6% from 1 × 103 CFU/mL to 1 × 108 CFU/mL), specific (compared to M. smegmatis mc2 155), and reusable (6 cycles).
treatment, S. epidermidis and M. smegmatis mc2 155 bacteria became purple and cyan in color, respectively. This may be caused by their different morphologies that generate a difference of reflection light. After treating the bacteria (108 CFU/mL) with FMSN-G7 (0.05 mg/mL) for 2 h, the purple color of S. epidermidis became transparent, and large aggregates were formed (Figure 3B). Without G7, no color change was observed when the bacteria were mixed with FMSN, and there was no obvious aggregate (Figure 3B). The same was true for M. smegmatis mc2 155 treated with FMSN-G7 (Figure 3B). These results indicate that FMSN-G7 acted as an effective platform for selectively capture and detection of S. epidermidis. The time frame for capture and detection can be modulated by the concentration of FMSN-G7 (Figure S10). When the particle concentration increased to 0.2 mg/mL, a rapid capture could be realized within 18 min. The capture kinetic of S. epidermidis (108 CFU/mL) by FMSN-G7 (0.05 mg/mL) was then studied. It can be seen clearly that this capture process is time-dependent (Figure 3C). After 2 h of treatment, the capture efficiency could be achieved to ∼99.3%. We further explored the possibility for capturing S. epidermidis at low concentrations. As shown in Figure 3D, after treating with FMSN-G7 (0.05 mg/mL) for 2 h, S. epidermidis at a concentration range from 1 × 103 CFU/mL to 1 × 108 CFU/mL can be captured with a efficiency of at least 98.6%. These preliminary results demonstrated that FMSN-G7 can act as an effective multivalent platform for capture and detection of S. epidermidis in a simple, fast and low-cost way. Another important point is that the FMSN-G7 is reusable. After treated with abundant free G7, FMSN-G7 from bacteria-bound FMSNG7 could be displaced and thus S. epidermidis bacteria could be
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ASSOCIATED CONTENT
S Supporting Information *
The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acsami.6b10989. Experimental details, materials characterization, and additional data (PDF)
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AUTHOR INFORMATION
Corresponding Authors
*E-mail:
[email protected]. *E-mail:
[email protected]. ORCID
Nanjing Hao: 0000-0003-3808-7941 Notes
The authors declare no competing financial interest. C
DOI: 10.1021/acsami.6b10989 ACS Appl. Mater. Interfaces XXXX, XXX, XXX−XXX
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ACS Applied Materials & Interfaces
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ACKNOWLEDGMENTS The authors thank the financial support from the National Science Foundation of China (81171454). Dr. Nanjing Hao thanks Prof. Mingdi Yan (University of Massachusetts Lowell, MA, U.S.A.) for her kindly discussion and revision of this manuscript. Dr. Nanjing Hao also thanks Prof. John Xiaojing Zhang for his support in Dartmouth College, NH, U.S.A.
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DOI: 10.1021/acsami.6b10989 ACS Appl. Mater. Interfaces XXXX, XXX, XXX−XXX
