Microscopic Observations of Superficial Ultrastructure of

Unworn Siloxane-Hydrogel Contact Lenses by Cryo-Scanning

Electron Microscopy
José M. González-Méijome,1 Antonio López-Alemany,2,3 José B. Almeida,1 Manuel A. Parafita,4
Miguel F. Refojo5
1
Department of Physics (Optometry), School of Sciences, University of Minho, Braga, Portugal

2
Ocular Surface, Cornea, and Contact Lens Research Group, Department of Optics, School of Optometry and Optics,
University of Valencia, Valencia, Spain

3
Institute of Ophthalmology and Optometry of La Costera (IOOC), Novetlé, Spain

4
Department of Surgery (Ophthalmology), School of Optics and Optometry, University of Santiago de Compostela, Santiago
de Compostela, Spain

5
Schepens Eye Research Institute, Harvard Medical School, Boston, Massachusetts

Received 31 March 2005; revised 12 May 2005; accepted 19 May 2005

Published online 23 September 2005 in Wiley InterScience (www.interscience.wiley.com). DOI: 10.1002/jbm.b.30386

Abstract: The purpose of this study was to analyze three commercial siloxane-hydrogel
contact lens materials, lotrafilcon A, balafilcon A, and galyfilcon A, by cryogenic scanning
electron microscopy (cryoSEM). The fully hydrated lenses were frozen in slush liquid nitrogen
and qualitatively observed in a cryogenic scanning electron microscope. The superficial
ultrastructure of the siloxane-hydrogels was observed at the areas where the lens fractured
during sample cryogenic preparation. There are qualitative differences among the three
examined materials in the complex polymer network structure existing between the outer
layer and the underlying polymer. CryoSEM, although destructive, is a useful tool to inves-
tigate the structure of polymers used in contact lenses. This technique allows the observation
of the inner structure of polymers in the hydrated state. The ultrastructure, the polymer
network underlying the outer surface of siloxane-hydrogels by cryoSEM microscopy, have
never been reported before. © 2005 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater
76B: 419 – 423, 2006

Keywords: microscopy; cryoSEM; silicone-hydrogel; siloxane-hydrogels; contact lens mate-

rials; polymer surface

INTRODUCTION and particularly cornea’s physiology.1 Minimum gas exchange

The advent of siloxane-hydrogel (Si-Hi) materials brought a
revolutionary advance to the contact lens field. Such materials
combine the comfort and hydrophilic nature of the conventional
hydrogels with the ability of siloxane moieties in the polymer to
transport oxygen and carbon dioxide. Gaseous exchange is nec-
essary for the maintenance of normal ocular surface homeostasis
Correspondence to: J. M. González-Méijome (e-mail: jgmeijome@fisica.uminho,pt)
Contract grant sponsor: Science and Technology Foundation (FCT), Ministry of
Science and Superior Education (MCES)
Contract grant sponsor: European Social Funding (to JMG-M); contract grant
number: 8281/2002
Present results have been presented in part at the ARVO 2005 Meeting, Fort
Lauderdale, FL, May 1–5, 2005. None of the authors has a financial or proprietary
interest in any material mentioned in this article.
© 2005 Wiley Periodicals, Inc.
criteria to avoid corneal compromise in the form of edema as a
primary result of hypoxia (decrease in oxygen) and hypercapnia
(increase in carbon dioxide) have been previously discussed.2
More recently, these criteria were revised by Harvitt and Bon-
anno.3 First-generation siloxane-hydrogel materials are repre-
sented by Focus Night & Day™ (lotrafilcon A; CIBA Vision,
Duluth, GA) and Purevision™ (balafilcon A; Bausch & Lomb,
Rochester, NY). Currently, two new materials have been deliv-
ered to the marketplace, which present some modifications from
those of the first generation. These new lenses are Acuvue
Advance™ (galyfilcon A; Vistakon, Jacksonville, FL) and
O2OPTIX (lotrafilcon B; CIBA Vision).
The surfaces of first-generation Si-Hi materials are hydro-
phobic because of the siloxane moieties migrating to their
surfaces and, hence, are nonwettable by tears and poorly
419
420 GONZÁLEZ-MÉIJOME ET AL.
TABLE I. Identification and Nominal Parameters of Contact Lenses Used in the Study
Acuvue Advance
Manufacturer Vistakon J&J
Material (USAN) galyfilcon A
FDA Group I I
Manufact. Procedure Cast molding
Surface Treatment None
Hydration 47%
Dk (barrer) 60
tolerated on the eye. Therefore, the lenses are finished with
treatments to obtain wettable surfaces: Purevision™ by
plasma oxidation4 and Focus Night & Day™ by plasma
polymerization of a mixture of trimethylsilane, oxygen, and
methane.5 Acuvue Advance™ does not need surface treat-
ment for good wetting according to the manufacturer. The
fourth lens, O2OPTIX, is the newest lens of this family
currently on the market, and no data about the superficial
structure or potential treatments to increase wettability are
available. This lens is not included in this study.
Several techniques have been applied to the microscopic
examination of biomaterials and, in particular, hydrogel con-
tact lens materials. X-ray photoelectron spectroscopy (XPS)
was used to investigate deposits on contact lenses.6 Sum-
frequency-generation (SFG) vibrational spectroscopy was
used to investigate contact lens surface and dehydration.7
Electrochemical impedance spectroscopy (EIS) was used to
investigate plasma coatings on siloxane-based hydrogels.5
Time-of-flight secondary ion mass spectrometry (ToF-SIMS)
was used to characterize the chemical surface of soft contact
lenses.8 Atomic force microscopy was applied to investigate
deposits,9 surface differences between soft contact lenses as a
function of the manufacturing process,10 and adhesion and
friction.7 Scanning electron microscopy (SEM) has contrib-
uted to the elucidation of deposit formation11,12 and the
porous structure of hydrogels.13
Cryogenic scanning electron microscopy (cryoSEM), also
known as low-temperature scanning electron microscopy
(LTSEM), has been applied to the investigation of the struc-
ture of hydrogels.14,15 This technique has also been named
aquaSEM by other authors, but the principles of sample
preparation and observation methodology seem to be equiv-
alent.16
CryoSEM is widely used to study biological samples,17
particularly in plant research.18,19 This technique requires that
the material be frozen in nitrogen before examination. In
hydrogels, this usually means the destruction of the material,
which is the main disadvantage of this technique. However,
the result of this process usually shows important features of
the material under study. CryoSEM has been applied recently
to the study of porous structure of “superporous hydro-
gels.”20,21 Of particular interest is the application of this
technique to the observation of the ultrastructure of polysac-
charide hydrogels.14
CryoSEM allows the observation of frozen biological
samples as well as hydrated polymer samples, in their natural
Focus Night & Day Purevision
Ciba Vision Bausch & Lomb
lotrafilcon A balafilcon A
III
Cast molding Cast molding
Plasma coating Plasma oxidation
24% 36%
140 99
physiological conditions without critical point dehydration as
required by conventional SEM.
In this study, we used cryoSEM to characterize unworn
siloxane-hydrogel contact lenses swollen in physiological
saline solution. To the best of our knowledge, this is the first
time that the structure of siloxane-hydrogel soft contact
lenses was studied by this technique. No images of new Si-Hi
materials are available in literature studied under this tech-
nique.
MATERIALS AND METHODS
First-generation siloxane-hydrogel contact lenses Focus
Night & Day™ (lotrafilcon A; CIBA Vision, Duluth, GA) and
Purevision™ (balafilcon A; Bausch & Lomb, Rochester, NY),
and newer siloxane-hydrogel contact lenses Acuvue Ad-
vance™ (galyfilcon A; Vistakon, Jacksonville, FL) are used in
this study. Technical details are summarized in Table I. The
lenses were obtained in the original containers filled with
physiological saline solutions.
Sample Preparation and CryoSEM Observations
The sample was frozen in slush N2 and attached to the
specimen holder of a CT-100C Cryo-transfer system (Oxford
Instruments, Oxford, UK) interfaced with a JEOL JSM-5410
scanning electron microscope (SEM). The sample was then
fractured and transferred from the cryostage to the micro-
scope sample stage, where the condensed surface water was
sublimed by controlled warming to ⫺90°C. Afterward, the
sample was transferred again to the cryostage to be coated by
gold using the sputtering procedure. Finally, the sample was
put back to the microscope sample stage to be examined at an
accelerating voltage of 15 KeV.
While frozen, the material suffers serious damage, which
induces the lens to break. However, these points of stress
resulted in good areas to observe not only the superficial but
also the underlying structure and bulk of the polymer.
RESULTS
Microphotographs in Figure 1 show the underlying ultrastruc-
ture of the three siloxane-hydrogel contact lenses. There are
significant differences between galyfilcon A and the other
 CRYOSEM ANALYSIS OF POLYMERS IN CONTACT LENSES 421

Figure 2. CryoSEM microphotographs of galyfilcon A. Microphoto-

graphs (c) and (d) correspond to details of image (b) at the marked
sites.

Microphotographs presented in Figure 5 highlight the ul-

trastructure linking the outer layer and the polymer bulk at
lower and higher magnification. These images show how
different the assembly pattern is in the three polymers. Bal-
afilcon A contact lens shows the tighter surface network
whose pores vary in size and density. Of particular relevance
is the round appearance adopted by the terminal ramifications
Figure 1. CryoSEM microphotographs at relatively low magnification
of the structure.
showing the zones where the outer layer collapsed showing the
underlying polymeric bulk and the ultrastructure between them.
DISCUSSION

two materials. In lotrafilcon A and balafilcon A, the ultra- CryoSEM allowed us to perform a morphological analysis of
structure appears as a loose network that attaches an outer the polymeric outer structure of hydrogels without dehydra-
membrane to the bulk of the polymeric network.
In Figure 2, the broken side of galyfilcon A contact lens
appears as a solid bulk overlying with a thin cover attached to
each other by a characteristic formation of lamellae with
small projections rounded at the end. High magnification
details in Figure 2(c,d) show relatively uniform surfaces with
a granulated appearance for the outer layer and polymeric
bulk.
In Figure 3, the ultrastructure underlying the outer layer of
lotrafilcon A displays a morphological pattern of porosity
where the pores are intercommunicated by a loose network of
filamentous structures. Note that Figure 3(a) is a magnifica-
tion of Figure 1(c,d) highlighting the inner side of the super-
ficial layer.
Figure 4 displays the superficial structure of balafilcon A.
Note that the outer surface of the superficial layer seen in
Figure 4(a,b) is similar to that presented in Figure 2(b). In
Figure 4(c), the surface and broken edge of the underlying
polymeric bulk is seen under higher magnification. In Figure
4(d), a macropore is evident under medium magnification. Figure 3. CryoSEM microphotographs of lotrafilcon A.
422 GONZÁLEZ-MÉIJOME ET AL.

The size of the pores seen in the balafilcon A material with

Figure 4. CryoSEM microphotographs of balafilcon A.
cryoSEM are larger that those observed with conventional
SEM.13 Differences in sample preparation between SEM and
cryoSEM probably account for such differences. In addition,
during freezing of the aqueous phase of the hydrogel, some
distension of the polymeric network could lead to enlarge-
ment of molecular spaces, increasing pore dimensions.
CryoSEM of frozen samples provides detailed information
about the structure, density, homogeneity, and arrangement
of the polymer network.14 The application of this technique to
contact lens polymers will be useful to detect changes in
surface and bulk structure as a consequence of use, or artifi-
cially created environments that could affect material prop-
erties such as changes in pH, osmolarity of solutions, tem-
perature, or interaction with biological fluids. CryoSEM
provides a powerful approach for direct imaging of macro-
molecular structures with high contrast and resolution of
hydrated samples.
We thank M. Planes and J. L. Moya (Polythecnic University of
Valencia) for their assistance with microscopy analysis.

tion. We have found a complex ultrastructure that links an

external membrane to the polymeric bulk of siloxane-hydro-
gel lenses never documented before. Although not identified,
it seems to be different for the three lenses studied, related to
the surface treatment of balafilcon A and lotrafilcon A to
improve wettability and to the presence of polyvinyl pyrro-
lidone (PVP) moieties on the surface of galafylcon A contact
lenses. The network or the ultrastructure observed under the
outer layer is denser for balafilcon A contact lens.
The second-generation siloxane hydrogel contact lens
Acuvue Advance ensures superficial hydration through an
internal wetting agent called Hydraclear™ which is derived
from the PVP family.22 Although significantly different from
the first-generation siloxane-hydrogels lenses, an ultrastruc-
ture underlying an outer thin layer is evident for this lens too.
The network or the ultrastructure observed under the outer
layer is denser for balafilcon A contact lens than the other two
lenses.
Pradny et al.16 obtained a similar structure for a macro-
porous hydrogel fabricated by crosslinking polymerization of
2-hydroxyethyl methacrylate (HEMA) with methacrylic acid
(MA) in the presence of NaCl particles. A similar appearance
was also observed on the matrix of alginate beads.14 These
authors concluded that such structures correspond to polysac-
charide chains and demonstrated that they were sensitive to
heat, becoming denser and tighter when exposed to elevated
temperatures. Within the field of hydrogel contact lenses,
microphotographs of the internal structure of conventional
hydrogels made of polymacon (poly-hydrodroxyethyl
methacrylate) (38% water content) and vasurfilcon A (copol-
ymer of methyl methacrylate and N-vinyl pyrrolidone, 74%
water content) revealed a similar structure.13 However, it
should be noted that images reported by Lopez-Alemany et
al.13 were taken with SEM at ⫻35,000 –⫻50,000 magnifica- Figure 5. Sequence of CryoSEM microphotographs comparing the
tion, much higher than that needed to observe the structures ultrastructure underlying the outer lens layer at different magnifica-
we presented here. tion.
 CRYOSEM ANALYSIS OF POLYMERS IN CONTACT LENSES
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