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Optics Express

Optics Express

  • Editor: Michael Duncan
  • Vol. 14, Iss. 19 — Sep. 18, 2006
  • pp: 8613–8621
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Freeform multiphoton excited microfabrication for biological applications using a rapid prototyping CAD-based approach

Lawrence P. Cunningham, Matthew P. Veilleux, and Paul J. Campagnola  »View Author Affiliations


Optics Express, Vol. 14, Issue 19, pp. 8613-8621 (2006)
http://dx.doi.org/10.1364/OE.14.008613


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Abstract

Multiphoton excited polymerization has attracted increasing attention as a powerful 3 dimensional nano/microfabrication tool. The nonlinear excitation confines the fabrication region to the focal volume allowing the potential to achieve freeform fabrication with submicron capabilities. We report the adaptation and use of a computer aided design (CAD) approach, based on rapid prototyping software, which exploits this potential for fabricating with protein and polymers in biologically compatible aqueous environments. 3D structures are drawn in the STL format creating a solid model that can be sliced, where the individual sections are then serially fabricated without overwriting previous layers. The method is shown for potential biological applications including microfluidics, cell entrapment, and tissue engineering

© 2006 Optical Society of America

1. Introduction

The main advantage in the MPE technique relative to other fabrication methods lies within its potential freeform capabilities and concurrent nano/microscale capabilities. A major challenge in the field has been implementing scanning methods to fully exploit this power. Many reports have used x-y-z stage scanning, as coupled with the appropriate control, allows user-defined fabrication in all 3 dimensions. However, this method is 2–3 orders of magnitude slower than laser-scanning through galvanometer mirrors. The latter is highly preferred for biological applications, where both the starting materials and resulting structure can readily suffer photodamage from prolonged exposure to the high peak powers required for efficient excitation. Additionally, we have previously shown that fabrication rate for proteins is 2–3 orders of magnitude slower than for polymers.[20

20. J. D. Pitts, P. J. Campagnola, G. A. Epling, and S. L. Goodman, “Reaction efficiencies for sub-micron multi-photon freeform fabrications of proteins and polymers with applications in sustained release,” Macromolecules 33, 1514–1523 (2000). [CrossRef]

] However since MPE fabrication is typically a threshold process, simply decreasing the pulse energy with longer pixel dwell times is not a tenable approach to this fabrication task. Thus repetitive laser galvo scanning with freeform capabilities would be the ideal approach.

While laser scanning with galvo mirrors provides great speed, these devices are not designed for complete random access to create user defined patterns. For example, the galvos and control software in laser scanning confocal microscopes are optimized for simple scanning patterns of lines and rectangles. To overcome this limitation, there have been several reports of approaches used to increase the flexibility of laser scanning for MPE fabrication. For example, we constructed an instrument that is capable of variety of geometrical patterns including polygons, ellipses, and circles, in both perimeter and solid patterns.[21

21. M. Sridhar, S. Basu, V. L. Scranton, and P. J. Campagnola, “Construction of a laser scanning microscope for multiphoton excited optical fabrication,” Rev. Sci. Instrum. 74, 3474–3477 (2003). [CrossRef]

] In an alternate approach, Braun [22

22. S. A. Pruzinsky and P. V. Braun, “Fabrication and characterization of two-photon polymerized features in colloidal crystals,” Adv. Funct. Mater. 15, 1995–2004 (2005). [CrossRef]

] and coworkers use modulated raster scanning, where by controlling the power during the scan with an electro-optic modulator, much greater flexibility was obtained. Maruo [23

23. S. Maruo, K. Ikuta, and H. Korogi, “Force-controllable, optically driven micromachines fabricated by single-step two-photon micro stereolithography,” J. Microelectromech. Syst. 12, 533–539 (2003). [CrossRef]

] utilized a combination of stage and circular laser scanning to construct an impressive range of micromachines including gears and actuators.

It is of great value for a range of biological applications to extend these ideas and utilize rapid galvo scanning to fabricate user-defined 3D objects directly from soluble proteins. In this article we describe our continuing efforts to achieve this goal, where we link rapid prototyping (RP) stereolithography software into our LabVIEW control code (described in Ref. [21

21. M. Sridhar, S. Basu, V. L. Scranton, and P. J. Campagnola, “Construction of a laser scanning microscope for multiphoton excited optical fabrication,” Rev. Sci. Instrum. 74, 3474–3477 (2003). [CrossRef]

]. RP utilizes the STL file format, which creates a triangular mesh of the surface of a solid to be fabricated. This method has been widely used in a variety of industries (e.g. automotive, aerospace, and medical device industries) to create macroscale (mm-cm) physical objects directly from CAD data sources. In this process any desired structure or combination of structures of user-defined complexity and intricacy can be drawn and then rapidly fabricated. Here we adapt this 3D fabrication method to biological applications by crosslinking proteins from a freely diffusing aqueous environment. The overall approach begins with 3D solid CAD drawing that is sliced into individual files, where these are fed to our laser scanning fabrication microscope. We show several examples of the method, including devices for cell sorting, cell encapsulation, and tissue engineering.

2. Experimental methods

2.1. Materials and methods

Octadecyltrichlorosilane (Gelest), anhydrous toluene (Aldrich), type I and type II collagen (Sigma), Bovine Serum Albumin (BSA), Rose Bengal (Sigma) were used as received. Water soluble ethoxylated trimethylolpropane triacrylate (TMPTA) was kindly provided by Sartomer.

2.2. Photochemistry

We have described the MPE photochemistry for crosslinking proteins and polymers in detail previously.[20

20. J. D. Pitts, P. J. Campagnola, G. A. Epling, and S. L. Goodman, “Reaction efficiencies for sub-micron multi-photon freeform fabrications of proteins and polymers with applications in sustained release,” Macromolecules 33, 1514–1523 (2000). [CrossRef]

] Briefly, we use two-photon excitation of Rose Bengal (1 mM) at 780 nm. Polymerization of the TMPTA requires a co-initiator, where 0.1M of TEA is used. Crosslinking proteins requires only the Rose Bengal photoactivator. BSA was used at a concentration of 10 mg/mL, or ~10-4M. Preparations were created on silanized microscope slides under cover slips that were separated from the slide using glass spacers, such that the enclosed volume was approximately 50 µL.

2.3. Fabrication instrument/microscope

The laser- scanning fabrication microscope has been described in detail [21

21. M. Sridhar, S. Basu, V. L. Scranton, and P. J. Campagnola, “Construction of a laser scanning microscope for multiphoton excited optical fabrication,” Rev. Sci. Instrum. 74, 3474–3477 (2003). [CrossRef]

] and is only briefly described here. The multi-photon excitation is achieved through a femtosecond near-infrared titanium-sapphire oscillator (Mira 900-F, Coherent) that is pumped by a 5 W Verdi Nd:YVO4 (Coherent). The laser is coupled to an upright microscope (Zeiss Axioskop,) that is equipped with both bright field and fluorescence optics, which are used as online diagnostics of the fabrication process. Scanning is achieved by a combination of the laser galvo scanning mirrors as well as a programmable x-y-z motorized stage. Typical pulse energies at the sample were approximately 500 pJ/pulse. A 20X, 0.75 numerical aperture (NA) objective lens was used for the fabrication. The code for operating this instrument for both the scan control and simultaneous data acquisition was written entirely using LabVIEW 7 (National Instruments) and is freely available on our website at:

http://www.cbit.uchc.edu/faculty_nv/campagnola/fabrication.html.

Two-photon fluorescence is used for imaging the fabricated structures, where the contrast arises from either residual Rose Bengal photoactivator or from staining with Rhodamine B or Di-4-ANEPPS. 3D stacks are acquired on an Olympus Fluoview modified for 2-photon imaging. Rendering of the stacks was performed with Imaris Bitplane.

2.4. CAD STL approach

Fig. 1. Flow chart of the possible options to create 2 and 3 dimension structures using STL files and LabVIEW.

3. Results

Here we demonstrate our CAD-based MPE fabrication in a microscopic aqueous environment by presenting several examples of biological applications using the STL format to fabricate objects from proteins and polymers. As a demonstrative example, we fabricated a microflow “tunnel” from a water soluble triacrylate. Solid renderings of four of the 20 STL slices are shown in Fig. 2(b). The resulting structure has dimensions of 130 µm (l) by 100 µm (w) by 90 µm (h), with wall thickness 10 µm. Following MPE polymerization, the chamber was washed with water three times to remove un-reacted monomer as well as any entrapped Rose Bengal photoactivator. The structure was stained with Rhodamine B and imaged by two-photon excited fluorescence (TPEF) at 830 nm. Figure 2 shows two views of the 3D rendered fluorescence image, where (b) and (c) show the two openings as well as the curved interior.

Fig. 2. MPE fabrication of a “tunnel”. The four structures in (a) show representative solid renderings of the steps of the process of creating the 3D structure. The images in (b) and (c) are two different projections of the 3D rendering of the Rhodamine B labeled structure.

This type of structure is not possible to create by lithographic methods or by conventional geometric laser scanning. It should be possible to use this fabrication technology to create microflow devices for sorting cells. As a rudimentary example, we have placed L1210 lymphocytes inside a MPE fabricated tunnel (325×140×172 µm). The structure was washed as described above. The cells were stained with the membrane staining dye Di-4-ANNEPS and introduced into the tunnel by microinjection, such that the cells flowed in a gentle vortex into the tunnel. A 3D TPEF stack was obtained and a one optical section near the bottom of the tunnel, containing the cells is shown in the left panel of Fig. 3, where the arrows point to the cells (diameter ~10 microns). The right panel shows an optical section near the middle region of the structure and cells are seen near the entrance. This type of structure could further be functionalized with cell attractive molecules such as fibronectin, collagen, or growth factors.

Fig. 3. TPEF optical sections of L1210 cells in the MPE fabricated microflow device, where (a) and (b) are near the bottom and top of the structure, respectively.

3.1. MPE crosslinked protein structures

While we demonstrated the CAD based control of the fabrication process with a water soluble acrylate, we ultimately wish to exploit the technology for biological applications where fabricated devices can be created from proteins. Here we show several simple examples to demonstrate the functionality of the approach. Figure 4(a) shows the 3D rendering of the TPEF image of a high aspect ratio cylinder fabricated from MPE crosslinked BSA, where the height is 60 µm, opening of 60 µm, and wall thickness of 1µm. This form of structure can be used to entrap cells to perform motility migration and studies. To this end as shown in Fig 4(b), we have a fabricated a cylinder from a mixture of crosslinked Texas Red labeled fibrinogen and BSA, where the former provides ECM cues through RGD binding sites,[24

24. J. Gailit, C. Clarke, D. Newman, M. G. Tonnesen, M. W. Mosesson, and R. A. Clark, “Human fibroblasts bind directly to fibrinogen at RGD sites through integrin alpha(v)beta3,” Exp Cell Res 232, 118–126 (1997). [CrossRef] [PubMed]

] and the latter is used to provide increased mechanical stability. The inner rectangular matrix was fabricated from the same mixture. The image in Fig. 4(b) shows two GFP expressing neurons (isolated from primary culture) that have migrated into the contained region and have attached to the inner matrix. A fibroblast at the right edge has also migrated towards this region. The general problems of cell motility and migration are not well understood and other work has reported the use of cell entrapment as a means to gain insight into these phenomena.[25

25. W.-G. Koh, A. Revzin, and M. V. Pishko, “Poly(ethylene glycol) Hydrogel microstructures encapsulating living cells,” Langmuir 18, 2459–2462 (2002). [CrossRef] [PubMed]

] We suggest that MPE fabricating the entrapment vessel as well functionalizing its interior may be a versatile tool to study motility and migration on attractive surfaces in a well-controlled, spatially confined environment.

Fig. 4. 3D dimensional cell-containment devices (a) is a high aspect ratio cylinder 60 microns tall with 1 micron thick walls MPE fabricated from BSA (b) Cylinder fabricated from crosslinked fibrinogen/BSA mixture is used to encompass primary neurons and fibroblasts. The inner scaffold is also fabricated from fibrinogen and the cells have migrated to this adhesive structure. Scale bar=50 microns.

Fig. 5. A tissue engineering scaffolds fabricated from fibrinogen and BSA., where (a) show a 3D fluorescence rendering of the structure, and (b) shows a GFP expressing fibroblast adhered to the scaffold 4 hours post-plating. Additional fibroblasts are seen to migrate toward the scaffold. Scale bar=25 microns.
Fig. 6. The fabrication process for the UCONN Oakleaf logo is shown where the CAD drawing, the TPE image, and 3d rendering are shown in the left, middle and right panels, respectively.

While the scaffold shown in Fig. 5 has more 3D structure than we have previously reported, native ECMs have significantly more complex topography. Our long-term goals are to use the freeform capabilities of MPE fabrication to replicate this structure. As a step in this direction, we can use the CAD control approach to fabricate complex, arbitrary 3D structures.

4. Discussion and conclusions

There have been other recent reports of using rapid prototyping software for tissue engineering applications.[26

26. Y. Lu, G. Mapili, G. Suhali, S. Chen, and K. Roy, “A digital micro-mirror device-based system for the microfabrication of complex, spatially patterned tissue engineering scaffolds,” J Biomed Mater Res A 77, 396–405 (2006). [PubMed]

, 27

27. M. N. Cooke, J. P. Fisher, D. Dean, C. Rimnac, and A. G. Mikos, “Use of stereolithography to manufacture critical-sized 3D biodegradable scaffolds for bone ingrowth,” J Biomed Mater Res B Appl Biomater 64B, 65–69 (2003). [CrossRef]

] However, these approaches have used either conventional commercially available STL machines, or direct light projection using standard components. Since these approaches are based on the standard methodologies, the resulting minimum feature sizes are in the range of a few hundred microns to several mm. By contrast the technology reported here adapts the powerful CAD based RP approach to the submicron and micron scale and performs the fabrication with biological materials in a biocompatible aqueous environment. This opens up the door to an array of applications in tissue engineering, biosensing, and cell biology.

Acknowledgments

We gratefully acknowledge support under an NIH 1U54-RR022232.

References and links

1.

W. Lee, S. A. Pruzinsky, and P. V. Braun, “Multi-photon polymerization of waveguide structures within three-dimensional photonic crystals,” Adv. Mater. 14, 271–274 (2002). [CrossRef]

2.

W. Zhou, S. M. Kuebler, K. L. Braun, T. Yu, J. K. Cammack, C. K. Ober, J. W. Perry, and S. R. Marder, “An efficient two-photon-generated photoacid applied to positive-tone 3D microfabrication,” Science 296, 1106–1109 (2002). [CrossRef] [PubMed]

3.

S. Kawata, H.-B. Sun, T. Tanaka, and K. Takada, “Microfabrication: finer features for functional microdevices,” Nature 412, 697–698 (2001). [CrossRef] [PubMed]

4.

S. Maruo, O. Nakamura, and S. Kawata, “Three-dimensional microfabrication with two-photon-absorbed photopolymerization,” Opt. Lett. 22, 132–134 (1997). [CrossRef] [PubMed]

5.

B. Kaehr, R. Allen, D. J. Javier, J. Currie, and J. B. Shear, “Guiding neuronal development with in situ microfabrication,” Proc. Natl. Acad. Sci. U S A 101, 16104–16108 (2004). [CrossRef] [PubMed]

6.

J. H. Strickler and W. W. Webb, “Three-dimensional optical data storage in refractive media by two-photon point excitation,” Opt. Lett. 16, 1780–1782 (1991). [CrossRef] [PubMed]

7.

C. N. LaFratta, T. Baldacchini, R. A. Farrer, J. T. Fourkas, M. C. Teich, B. E. A. Saleh, and M. J. Naughton, “Replication of two-photon-polymerized structures with extremely high aspect ratios and large overhangs,” J. Phys. Chem. B 108, 11256–11258 (2004). [CrossRef]

8.

F. J. Qi, Y. Li, H. C. Guo, H. Yang, and Q. H. Gong, “Wavy lines in two-photon photopolymerization microfabrication,” Opt. Express 12, 4725–4730 (2004). [CrossRef] [PubMed]

9.

T. Baldacchini, A. C. Pons, J. Pons, C. N. LaFratta, J. T. Fourkas, Y. Sun, and M. J. Naughton, “Multiphoton laser direct writing of two-dimensional silver structures,” Opt. Express 13, 1275–1280 (2005). [CrossRef] [PubMed]

10.

J. Serbin, A. Egbert, A. Ostendorf, B. N. Chichkov, R. Houbertz, G. Domann, J. Schulz, C. Cronauer, L. Frhlich, and M. Popall, “Femtosecond laser-induced two-photon polymerization of inorganic organic hybrid materials for applications in photonics,” Opt. Lett. 28, 301–303 (2003). [CrossRef] [PubMed]

11.

J. Serbin and M. Gu, “Superprism phenomena in waveguide-coupled woodpile structures fabricated by two-photon polymerization,” Opt. Express 14, 3563–3568 (2006). [CrossRef] [PubMed]

12.

J. Serbin, A. Ovsianikov, and B. Chichkov, “Fabrication of woodpile structures by two-photon polymerization and investigation of their optical properties,” Opt. Express 12, 5221–5228 (2004). [CrossRef] [PubMed]

13.

A. Doraiswamy, C. Jin, R. J. Narayan, P. Mageswaran, P. Mente, R. Modi, R. Auyeung, D. B. Chrisey, A. Ovsianikov, and B. Chichkov, “Two photon induced polymerization of organic-inorganic hybrid biomaterials for microstructured medical devices,” Acta Biomaterialia 2, 267–275 (2006). [CrossRef] [PubMed]

14.

A. Fujita, K. Fujita, O. Nakamura, T. Matsuda, and S. Kawata, “Control of cardiomyocyte orientation on a microscaffold fabricated by photopolymerization with laser beam interference,” J Biomed Opt 11, 021015 (2006). [CrossRef] [PubMed]

15.

S. Basu, C. W. Wolgemuth, and P. J. Campagnola, “Measurement of normal and anomalous diffusion of dyes within protein structures fabricated via multi-photon excited crosslinking,” Biomacromolecules 5, 2347–2357 (2004). [CrossRef] [PubMed]

16.

S. Basu and P. J. Campagnola, “Enzymatic activity of alkaline phosphatase inside protein and polymer structures fabricated via multi-photon excitation,” Biomacromolecules 5, 572–579 (2004). [CrossRef] [PubMed]

17.

S. Basu and P. J. Campagnola, “Properties of crosslinked protein matrices for tissue engineering applications synthesized by multiphoton excitation,” J Biomed Mater Res 71A, 359–368 (2004). [CrossRef]

18.

S. Basu, L. P. Cunningham, G. Pins, K. Bush, R. Toboada, A. R. Howell, J. Wang, and P. J. Campagnola, “Multi-photon excited fabrication of collagen matrices crosslinked by a modified benzophenone dimer: bioactivity and enzymatic degradation,” Biomacromolecules 6, 1465–1474 (2005). [CrossRef] [PubMed]

19.

G. D. Pins, K. A. Bush, L. P. Cunningham, and P. J. Campagnola, “Multiphoton excited fabricated nano and micropatterned extracellular matrix proteins direct cellular morphology,” J. Biomed. Mat. Res. 78A, 194–204 (2006). [CrossRef]

20.

J. D. Pitts, P. J. Campagnola, G. A. Epling, and S. L. Goodman, “Reaction efficiencies for sub-micron multi-photon freeform fabrications of proteins and polymers with applications in sustained release,” Macromolecules 33, 1514–1523 (2000). [CrossRef]

21.

M. Sridhar, S. Basu, V. L. Scranton, and P. J. Campagnola, “Construction of a laser scanning microscope for multiphoton excited optical fabrication,” Rev. Sci. Instrum. 74, 3474–3477 (2003). [CrossRef]

22.

S. A. Pruzinsky and P. V. Braun, “Fabrication and characterization of two-photon polymerized features in colloidal crystals,” Adv. Funct. Mater. 15, 1995–2004 (2005). [CrossRef]

23.

S. Maruo, K. Ikuta, and H. Korogi, “Force-controllable, optically driven micromachines fabricated by single-step two-photon micro stereolithography,” J. Microelectromech. Syst. 12, 533–539 (2003). [CrossRef]

24.

J. Gailit, C. Clarke, D. Newman, M. G. Tonnesen, M. W. Mosesson, and R. A. Clark, “Human fibroblasts bind directly to fibrinogen at RGD sites through integrin alpha(v)beta3,” Exp Cell Res 232, 118–126 (1997). [CrossRef] [PubMed]

25.

W.-G. Koh, A. Revzin, and M. V. Pishko, “Poly(ethylene glycol) Hydrogel microstructures encapsulating living cells,” Langmuir 18, 2459–2462 (2002). [CrossRef] [PubMed]

26.

Y. Lu, G. Mapili, G. Suhali, S. Chen, and K. Roy, “A digital micro-mirror device-based system for the microfabrication of complex, spatially patterned tissue engineering scaffolds,” J Biomed Mater Res A 77, 396–405 (2006). [PubMed]

27.

M. N. Cooke, J. P. Fisher, D. Dean, C. Rimnac, and A. G. Mikos, “Use of stereolithography to manufacture critical-sized 3D biodegradable scaffolds for bone ingrowth,” J Biomed Mater Res B Appl Biomater 64B, 65–69 (2003). [CrossRef]

OCIS Codes
(120.4610) Instrumentation, measurement, and metrology : Optical fabrication
(180.6900) Microscopy : Three-dimensional microscopy
(190.4180) Nonlinear optics : Multiphoton processes
(220.4000) Optical design and fabrication : Microstructure fabrication
(350.3450) Other areas of optics : Laser-induced chemistry

ToC Category:
Instrumentation, Measurement, and Metrology

History
Original Manuscript: April 28, 2006
Revised Manuscript: July 19, 2006
Manuscript Accepted: August 11, 2006
Published: September 18, 2006

Virtual Issues
Vol. 1, Iss. 10 Virtual Journal for Biomedical Optics

Citation
Lawrence P. Cunningham, Matthew P. Veilleux, and Paul J. Campagnola, "Freeform multiphoton excited microfabrication for biological applications using a rapid prototyping CAD-based approach," Opt. Express 14, 8613-8621 (2006)
http://www.opticsinfobase.org/oe/abstract.cfm?URI=oe-14-19-8613


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References

  1. W. Lee, S. A. Pruzinsky, and P. V. Braun, "Multi-photon polymerization of waveguide structures within three-dimensional photonic crystals," Adv. Mater. 14, 271-274 (2002). [CrossRef]
  2. W. Zhou, S. M. Kuebler, K. L. Braun, T. Yu, J. K. Cammack, C. K. Ober, J. W. Perry, and S. R. Marder, "An efficient two-photon-generated photoacid applied to positive-tone 3D microfabrication," Science 296, 1106-1109 (2002). [CrossRef] [PubMed]
  3. S. Kawata, H.-B. Sun, T. Tanaka, and K. Takada, "Microfabrication: finer features for functional microdevices," Nature 412, 697-698 (2001). [CrossRef] [PubMed]
  4. S. Maruo, O. Nakamura, and S. Kawata, "Three-dimensional microfabrication with two-photon-absorbed photopolymerization," Opt. Lett. 22, 132-134 (1997). [CrossRef] [PubMed]
  5. B. Kaehr, R. Allen, D. J. Javier, J. Currie, and J. B. Shear, "Guiding neuronal development with in situ microfabrication," Proc. Natl. Acad. Sci. U S A 101, 16104-16108 (2004). [CrossRef] [PubMed]
  6. J. H. Strickler and W. W. Webb, "Three-dimensional optical data storage in refractive media by two-photon point excitation," Opt. Lett. 16, 1780-1782 (1991). [CrossRef] [PubMed]
  7. C. N. LaFratta, T. Baldacchini, R. A. Farrer, J. T. Fourkas, M. C. Teich, B. E. A. Saleh, and M. J. Naughton, "Replication of two-photon-polymerized structures with extremely high aspect ratios and large overhangs," J. Phys. Chem. B 108, 11256 - 11258 (2004). [CrossRef]
  8. F. J. Qi, Y. Li, H. C. Guo, H. Yang, and Q. H. Gong, "Wavy lines in two-photon photopolymerization microfabrication," Opt. Express 12, 4725-4730 (2004). [CrossRef] [PubMed]
  9. T. Baldacchini, A. C. Pons, J. Pons, C. N. LaFratta, J. T. Fourkas, Y. Sun, and M. J. Naughton, "Multiphoton laser direct writing of two-dimensional silver structures," Opt. Express 13, 1275-1280 (2005). [CrossRef] [PubMed]
  10. J. Serbin, A. Egbert, A. Ostendorf, B. N. Chichkov, R. Houbertz, G. Domann, J. Schulz, C. Cronauer, L. Frhlich, and M. Popall, "Femtosecond laser-induced two-photon polymerization of inorganic organic hybrid materials for applications in photonics," Opt. Lett. 28, 301-303 (2003). [CrossRef] [PubMed]
  11. J. Serbin and M. Gu, "Superprism phenomena in waveguide-coupled woodpile structures fabricated by two-photon polymerization," Opt. Express 14, 3563-3568 (2006). [CrossRef] [PubMed]
  12. J. Serbin, A. Ovsianikov, and B. Chichkov, "Fabrication of woodpile structures by two-photon polymerization and investigation of their optical properties," Opt. Express 12, 5221-5228 (2004). [CrossRef] [PubMed]
  13. A. Doraiswamy, C. Jin, R. J. Narayan, P. Mageswaran, P. Mente, R. Modi, R. Auyeung, D. B. Chrisey, A. Ovsianikov, and B. Chichkov, "Two photon induced polymerization of organic-inorganic hybrid biomaterials for microstructured medical devices," Acta Biomaterialia 2, 267-275 (2006). [CrossRef] [PubMed]
  14. A. Fujita, K. Fujita, O. Nakamura, T. Matsuda, and S. Kawata, "Control of cardiomyocyte orientation on a microscaffold fabricated by photopolymerization with laser beam interference," J Biomed Opt 11, 021015 (2006). [CrossRef] [PubMed]
  15. S. Basu, C. W. Wolgemuth, and P. J. Campagnola, "Measurement of normal and anomalous diffusion of dyes within protein structures fabricated via multi-photon excited crosslinking," Biomacromolecules 5, 2347-2357 (2004). [CrossRef] [PubMed]
  16. S. Basu and P. J. Campagnola, "Enzymatic activity of alkaline phosphatase inside protein and polymer structures fabricated via multi-photon excitation," Biomacromolecules 5, 572-579 (2004). [CrossRef] [PubMed]
  17. S. Basu and P. J. Campagnola, "Properties of crosslinked protein matrices for tissue engineering applications synthesized by multiphoton excitation," J Biomed Mater Res 71A, 359-368 (2004). [CrossRef]
  18. S. Basu, L. P. Cunningham, G. Pins, K. Bush, R. Toboada, A. R. Howell, J. Wang, and P. J. Campagnola, "Multi-photon excited fabrication of collagen matrices crosslinked by a modified benzophenone dimer: bioactivity and enzymatic degradation," Biomacromolecules 6, 1465-1474 (2005). [CrossRef] [PubMed]
  19. G. D. Pins, K. A. Bush, L. P. Cunningham, and P. J. Campagnola, "Multiphoton excited fabricated nano and micropatterned extracellular matrix proteins direct cellular morphology," J. Biomed. Mat. Res. 78A, 194-204 (2006). [CrossRef]
  20. J. D. Pitts, P. J. Campagnola, G. A. Epling, and S. L. Goodman, "Reaction efficiencies for sub-micron multi-photon freeform fabrications of proteins and polymers with applications in sustained release," Macromolecules 33, 1514-1523 (2000). [CrossRef]
  21. M. Sridhar, S. Basu, V. L. Scranton, and P. J. Campagnola, "Construction of a laser scanning microscope for multiphoton excited optical fabrication," Rev. Sci. Instrum. 74, 3474-3477 (2003). [CrossRef]
  22. S. A. Pruzinsky and P. V. Braun, "Fabrication and characterization of two-photon polymerized features in colloidal crystals," Adv. Funct. Mater. 15, 1995-2004 (2005). [CrossRef]
  23. S. Maruo, K. Ikuta, and H. Korogi, "Force-controllable, optically driven micromachines fabricated by single-step two-photon micro stereolithography," J. Microelectromech. Syst. 12, 533-539 (2003). [CrossRef]
  24. J. Gailit, C. Clarke, D. Newman, M. G. Tonnesen, M. W. Mosesson, and R. A. Clark, "Human fibroblasts bind directly to fibrinogen at RGD sites through integrin alpha(v)beta3," Exp Cell Res 232, 118-126 (1997). [CrossRef] [PubMed]
  25. W.-G. Koh, A. Revzin, and M. V. Pishko, "Poly(ethylene glycol) Hydrogel microstructures encapsulating living cells," Langmuir 18, 2459-2462 (2002). [CrossRef] [PubMed]
  26. Y. Lu, G. Mapili, G. Suhali, S. Chen, and K. Roy, "A digital micro-mirror device-based system for the microfabrication of complex, spatially patterned tissue engineering scaffolds," J Biomed Mater Res A 77, 396-405 (2006). [PubMed]
  27. M. N. Cooke, J. P. Fisher, D. Dean, C. Rimnac, and A. G. Mikos, "Use of stereolithography to manufacture critical-sized 3D biodegradable scaffolds for bone ingrowth," J Biomed Mater Res B Appl Biomater 64B, 65-69 (2003). [CrossRef]

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