Spring 2005 Senior Design Project

Rachel E. Miller - Dept. of Bioengineering
The Pennsylvania State University, University Park, PA 16802

Sponsor: Dr. Christopher Siedlecki, Cardiovascular Biomaterials Lab
The Pennsylvania State University, Hershey, PA 17033

Executive Summary:

Biomedical devices, ranging from catheters to ventricular assist devices, are used by the millions annually, and in 2002, cardiovascular device sales in the United States totaled $6 billion [1]. Many novel coatings and materials have been developed for these applications, yet the ideal materials for these varied uses remain to be discovered [2-5]. An important strategic thrust in human healthcare is nanomedicine, as articulated by the National Institutes of Health (NIH) roadmap. As applied to biomaterials, nanomedicine implies the use of molecular self-assembly as the next step in the development of novel biocompatible materials. In this work, the novel thin films were used in order to design a coating for glass and silicon surfaces that is less thrombogenic than currently available thin film coatings. Material surfaces were engineered to have nanoscopic regions (“islands”) of one chemical functionality surrounded by a different chemical functionality. The surface was verified by the use of atomic force microscopy (AFM) (Fig. 1) and surface energy measurements (contact angle tensiometry). A flow device (Fig. 2) was designed and implemented to apply this coating to ~500 µm diameter glass beads. The new coating was evaluated and compared to other thin film coatings through the use of these glass beads in a previously published human blood plasma (the non-cellular portion of blood) coagulation assay [6, 7] and mathematical model [8]. The results indicate that the thin films bearing islands produced longer human blood plasma coagulation times than uniform thin films, indicating that surface engineering on the nanoscale may lead to more hemocompatible materials.

Fig. 1. AFM images in sets of 3, from left to right: height mode, friction mode trace, friction mode retrace. A) APS islands on glass substrates after a deposition time of 3 min at 4°C. B) APS islands back-filled with BTS with an island diameter of approximately 400 nm.

Fig. 2. Experimental set-up used for deposition of islands onto glass beads.

 

Download Final Report in Microsoft Word format: Final Report.doc

A summary of this report is provided below.

Overview:

  • 2002 – U.S. cardiovascular sales totaled $6 billion [1].
  • Primary design criterion = non-thrombogenic surface (one that does not promote the formation of clots)
  • Advances made in minimizing clots in cardiovascular devices, but some risk remains [1-5].
  • Nanomedicine is being emphasized as a new pathway to discovery by the National Institutes of Health (NIH) [6].
  • Nanomedicine implies the use of molecular self-assembly as the next step in the development of novel biocompatible materials.

Design Criteria:

  • Islands of one chemical functionality that cover 10-30% of the total surface area of a 12 mm diameter glass coverslip and should be no greater than 1 µm in diameter.
  • Islands will be surrounded by a different chemical functionality.
  • Device designed and constructed in order to apply coating to 500 µm diameter glass beads.
  • Beads will be used in blood coagulation assay to compare the hemocompatibility of this coating to previously developed coatings.
  • Data obtained from these assays will be quantified through a mathematical model that outputs a “catalytic potential” indicating the amount that the material activates blood coagulation.
  • Thin film must be able to be formed in under three hours using standard laboratory equipment.
  • Thin film should result in a smooth surface with a root-mean-square roughness less than 0.3 nm.
Design Alternatives:

Thin Film Creation on Glass Coverslips

Option 1: Use APS for island formation and surround the islands with BTS. Use a concentration of 0.01% by volume APS and 0.1% by volume BTS at room temperature.

Option 2: Use APS for island formation and surround the islands with BTS. Use a concentration of 0.0001% by volume APS and 0.1% by volume BTS at 4°C (temperature inside commercial refrigerator) in order to slow the reaction kinetics and exert tighter control over the deposition process.

Option 3: Use 4-aminobutyltriethoxysilane (ABS) for island formation and surround the islands with BTS. Use a concentration of 0.01% by volume ABS and 0.1% by volume BTS at room temperature.

Island Coating Device for Glass Beads

Option 1: Coat beads by using a method similar to the method used to coat glass coverslips. This would involve placing the beads in a petri dish, incubating in chemical solution, and aspirating the solution to end chemical deposition.

Option 2: Flow chemical solution over beads using the experimental set-up depicted in Fig 2.

Evaluation:

Coverslip surfaces were analyzed by:
  • Imaging with Nanoscope IIIa Multimode AFM (Digital Instruments, Inc. Santa Barbara, CA)
  • Measuring surface energy with Contact Angle Measuring System G10 (KRÜSS GmbH, Hamburg, Germany)
  • Calculating surface area coverage of the islands with MATLAB® (The MathWorks, Inc. Natick, MA) program.
  • See Fig. 1 and Table 1.
Table 1. APS Islands Back-filled with BTS at 4°C
 
Deposition Times (min)
 
3
4
5
Horizontal Static Contact Angle
(mean±st. dev.) (°)
97 ± 1
97 ± 1
97 ± 1
RMS Roughness (nm)
0.16 ± 0.02
0.159 ± 0.006
0.17 ± 0.02
Island Surface Area Coverage (%)
12.6 ± 1.2
7.0 ± 3.2
8.0 ± 0.4

 

Beads were evaluated for hemocompatibility by:

  • Previously published in vitro whole human blood plasma coagulation assay and mathematical model [7-9].
  • See Fig. 3 and Table 2.

Fig. 3. Surface area titrations with glass beads of varying surface energy plotting observed coagulation time as a function of procoagulant surface area per mL test solution. Note that “Nano APS/BTS” indicates beads with nanoscale chemical heterogeneity with approximately 10% of the bead surface coated in APS and 90% covered by BTS. The labels “50 OTS/50 APS” and “90 OTS/10 APS” indicate physical mixtures of beads bearing two different, uniform silane chemistries in the indicated percentages.

Table 2. Procoagulant Catalytic Potentials

Procoagulant Advancing/Receding Contact Angle (°)

Ksat (mL/m^2)
N = 3

Clean Glass 0/0 (5.3 ± 1.4) x 10^2
APS 70/40 1.6 ± 0.2
OTS 110/90 0.6 ± 0.1
BTS 93/80 0.7 ± 0.1
Nano APS/BTS 93/81 0.03 ± 0.01
90 OTS/10 APS -- 0.7 ± 0.1
50 OTS/50 APS -- 1.0 ± 0.08

Budget:

The total cost for the production of these glass coverslips and beads is the sum of materials, labor, and overhead. This project was completed by one person over a span of 10 weeks. Based on an estimate of working 12 hours per week at a salary of $10 per hour, the labor costs would have totaled $1200. As listed in Table 4.1, the materials needed cost a total of $1738. In addition, overhead costs can be estimated to be approximately $50 per week for a total of $500. Therefore, the total amount that would be spent for the production of these coatings on a small scale is $3438. Since there is currently no reason for these coatings to be produced for the general public, a cost of large-scale manufacturing will not be given.

Deliverables:

  • Protocols and experimental set-ups for developing thin films of two different chemical functionalities on flat and curved surfaces.
  • Glass coverslips and glass beads coated with a novel thin film that is more hemocompatible than currently available SAMs.

References:

[1] B. D. Ratner, A. S. Hoffman, F. J. Schoen, and J. E. Lemons, Biomaterials Science, 2nd ed. New York: Elsevier Academic Press, 2004.

[2] N. Nakabayashi and D. F. Williams, "Preparation of non-thrombogenic materials using 2-methacryloyloxyethyl phosphorylcholine," Biomaterials, vol. 24, pp. 2431-2435, 2003.

[3] B. D. Ratner, "Blood Compatibility - A Perspective," J. Biomat. Sci. Polym. Ed., vol. 11, pp. 1107-1119, 2000.

[4] J. L. Brash, "Exploiting the current paradigm of blood-material interactions for the rational design of blood-compatible materials.," J. Biomat. Sci. Polym. Ed., vol. 11, pp. 1135-1146, 2000.

[5] M. B. Gorbet and M. V. Sefton, "Biomaterial-associated thrombosis: roles of coagulation factors, complement, platelets and leukocytes," Biomaterials, vol. 25, pp. 5681-5703, 2004.

[6] E. Zerhouni, “The NIH Roadmap,” Science, vol. 302, pp.63-64, 72, 2003.

[7] E. A. Vogler, J. C. Graper, G. R. Harper, L. M. Lander, and W. J. Brittain, "Contact Activation of the Plasma Coagulation Cascade.," J. Biomed. Mat. Res., vol. 29, pp. 1005-1016, 1995.

[8] E. A. Vogler, J. G. Nadeau, and J. C. Graper, "Contact Activation of the Plasma Coagulation Cascade. 3. Biophysical Aspects of Thrombin Binding Anticoagulants.," J. Biomed. Mat. Res., vol. 40, pp. 92-103, 1997.

[9] R. Zhuo, R. Miller, K. Bussard, C. A. Siedlecki, and E. A. Vogler, "Procoagulant Stimulus Processing by the Intrinsic Pathway of Blood Plasma Coagulation," Biomaterials, vol. 26, pp. 2965-2973, 2005.