Centre for Contact Lens Research

Purpose: To determine the impact of incubation solution composition on protein deposition to silicone hydrogel (SH) contact lenses using a simplistic and a complex model of the tear film. Methods: Three SH materials - senofilcon A (SA), lotrafilcon B (LB), and balafilcon A (BA) - were incubated in two different solutions; Solution A was a simplistic augmented buffered saline solution containing a single protein, whereas Solution B was a complex artificial tear solution (ATS), containing the augmented buffered saline solution in addition to proteins, lipids, and mucins (pH=7.4). The proteins of interest (lysozyme, lactoferrin, albumin) were radiolabeled with Iodine-125 (2% protein of interest) and the accumulation of the conjugated protein to the lens materials was determined after 1, 7, 14, and 28 days of incubation. Protein deposition was measured using a gamma counter and the raw data were translated into absolute amounts (μg/lens) via extrapolation from standards. Results: After 28 days, lysozyme uptake was significantly lower on BA lenses when incubated in Solution A (33.7 μg) compared to Solution B (56.2 μg), p0.05. LB lenses also deposited similar amounts of lysozyme for both solutions (Solution A: 5.0 μg, Solution B: 4.7 μg, p>0.05). After 28 days, BA lenses accumulated approximately twice the amount of lactoferrin than the other lens materials, with 30.3 μg depositing when exposed to Solution A and 22.0 μg with Solution B. The difference between the two solutions was statistically significant (p0.05. LB lenses also deposited similar amounts of lysozyme for both solutions (Solution A: 5.0 μg, Solution B: 4.7 μg, p>0.05). After 28 days, BA lenses accumulated approximately twice the amount of lactoferrin than the other lens materials, with 30.3 μg depositing when exposed to Solution A and 22.0 μg with Solution B. The difference between the two solutions was statistically significant (p0.05). After 28 days, albumin deposition onto BA lenses was significantly greater when lenses were incubated in Solution B (1.7 μg) compared to Solution A (0.9 μg), p0.05). LB lenses incubated in Solution A deposited more albumin compared to Solution B (0.9 μg versus 0.6 μg), p=0.003. Discussion: Protein deposition onto SH materials varied when contact lenses were incubated in either a complex ATS compared to a single protein solution. More lysozyme accumulated onto BA lenses incubated in a complex analog of the human tear film, whereas lactoferrin deposited onto SA lenses independent of incubation solution composition. To better mimic the ex vivo environment, future studies should use more appropriate analogs of the tear film. © 2012 Molecular Vision.

Proteins are a key component in body fluids and adhere to most biomaterials within seconds of their exposure. The tear film consists of more than >400 different proteins, ranging in size from 10 to 2360 kDa, with a net charge of pH 1-11. Protein deposition rates on poly-2-hydroxyethyl methacrylate (pHEMA) and silicone hydrogel soft contact lenses have been determined using a number of ex vivo and in vitro experiments. Ionic, high water pHEMA-based lenses attract the highest amount of tear film protein (1300 μg/lens), due to an electrostatic attraction between the material and positively charged lysozyme. All other types of pHEMA-based lenses deposit typically less than 100 μg/lens. Silicone hydrogel lenses attract less protein than pHEMA-based materials, with <10 μg/lens for non-ionic and up to 34 μg/lens for ionic materials. Despite the low protein rates on silicone hydrogel lenses, the percentage of denatured protein is typically higher than that seen on pHEMA-based lenses. Newer approaches incorporating phosphorylcholine, polyethers or hyaluronic acid into potential contact lens materials result in reduced protein deposition rates compared to current lens materials. © 2012 British Contact Lens Association.

Purpose. This non-dispensing cross-over study was conducted to determine if lenses presoaked in Opti-Free RepleniSH (OFR) or ReNu MultiPlus (RMP) cause solution-induced corneal staining (SICS) and subsequent cell sloughing before the typical 2 h in vivo examination point. Methods. Study lenses (PureVision) were worn bilaterally by 13 participants for periods of 15, 30, 60, and 120 min using two different contralateral care regimen pairings. The lens worn on the test eye was soaked overnight in either OFR or RMP and the control eye in Clear Care (CC). After lens removal, corneal staining was rated on a scale of 0 (negligible) to 100 (severe) for four peripheral quadrants and the central region, and the differential global staining score was calculated by subtracting baseline staining scores. Following the staining assessment, corneal cells were collected from the ocular surface using a non-contact irrigation system to determine ocular cell shedding rates. Results. Differential global staining score with OFR was greater than CC with the differences being statistically significant at 30 and 60 min (p 0.05). Conclusions. SICS occurred earlier but to a significantly lower degree when PureVision lenses were presoaked in OFR compared with RMP, while lenses presoaked in CC did not cause SICS. Ocular surface cell shedding after lens removal was not impacted by lens wear durations of ≤2 h. © 2012 American Academy of Optometry.

PURPOSE: To investigate the impact of lactoferrin and lipids on the kinetic deposition of lysozyme on silicone and conventional hydrogel lenses, using a complex artificial tear solution (ATS). METHODS: Two silicone hydrogel lenses (AIR OPTIX AQUA; lotrafilcon B and ACUVUE OASYS; senofilcon A) and two conventional hydrogel lenses (ACUVUE 2; etafilcon A and PROCLEAR; omafilcon A) were investigated. Lenses were incubated in four different solutions: a complex ATS consisting of various salts, lipids, proteins, and mucins, an ATS without lactoferrin (ATS w/o Lac), an ATS without lipids (ATS w/o Lip), and an ATS without lactoferrin and lipids (ATS w/o Lac & Lip), each containing 2% radiolabeled (125I) lysozyme (1.9 mg/ml). After each time point (4, 12 h and 1, 2, 3, 5, 7, 14, 21, 28 days), the amount of lysozyme per lens was quantified. RESULTS: After 28 days, lotrafilcon B lenses incubated in ATS deposited significantly less lysozyme (9.7 ± 1.4 μg) than when incubated in solutions not containing lactoferrin and lipids (more than 11.8 μg) (p < 0.001). Lysozyme uptake to senofilcon A lenses was higher in ATS w/o Lip (5.3 ± 0.1 μg) compared with other solutions (less than 3.9 μg) (p < 0.001). Etafilcon A lenses deposited the most lysozyme in all four solutions compared with the rest of the lens types (p < 0.001). For etafilcon A lenses, less lysozyme was deposited when incubated in ATS w/o Lip (588.6 ± 0.4 μg) compared with the other solutions (more than 642.6 μg) (p < 0.001). Omafilcon A lenses in ATS w/o Lac accumulated significantly less lysozyme (12.8 ± 1.0 μg) compared with the other solutions (more than 14.2 μg) (p < 0.001). CONCLUSIONS: An ATS containing lactoferrin and lipids impacts lysozyme deposition on both silicone and conventional hydrogel contact lenses. When performing in vitro experiments to study protein deposition on contact lenses, more complex models should be used to better mimic the human tear film.

Model silicone hydrogel contact lenses, comprised of N,N-dimethylacrylamide and methacryloxypropyltris (trimethylsiloxy) silane, were fabricated and hyaluronic acid (HA) was incorporated as an internal wetting agent using a dendrimer-based method. HA and dendrimers were loaded into the silicone hydrogels and cross-linked using 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide chemistry. The presence and location of HA in the hydrogels was confirmed using X-ray photoelectron spectroscopy and confocal laser scanning microscopy, respectively. The effects of the presence of HA on the silicone hydrogels on hydrophilicity, swelling behavior, transparency, and lysozyme sorption and denaturation were evaluated. The results showed that HA increased the hydrophilicity and the equilibrium water content of the hydrogels without affecting transparency. HA also significantly decreased the amount of lysozyme sorption (p < 0.002). HA had no effect on lysozyme denaturation in hydrogels containing 0% and 1.7% methacrylic acid (MAA) (by weight) but when the amount of MAA was increased to 5%, the level of lysozyme denaturation was significantly lower compared to control materials. These results suggest that HA has great potential to be used as a wetting agent in silicone hydrogel contact lenses to improve wettability and to decrease lysozyme sorption and denaturation.

Purpose: To investigate the efficiency of lysozyme and albumin removal from silicone hydrogel and conventional contact lenses, using a polyhexamethylene biguanide multipurpose solution (MPS) in a soaking or rubbing/soaking application and a hydrogen peroxide system (H2O2). Methods: Etafilcon A, lotrafilcon B and balafilcon A materials were incubated in protein solutions for up to 14 days. Lenses were either placed in radiolabeled protein to quantify the amount deposited or in fluorescent-conjugated protein to identify its location, using confocal laser scanning microscopy (CLSM). Lenses were either rinsed with PBS or soaked overnight in H2O2 or MPS with and without lens rubbing. Results: After 14 days lysozyme was highest on etafilcon A (2,200 μg) >balafilcon A (50 μg) >lotrafilcon B (9.7 μg) and albumin was highest on balafilcon A (1.9 μg) =lotrafilcon B (1.8 μg) >etafilcon A (0.2 μg). Lysozyme removal was greatest for balafilcon A >etafilcon A >lotrafilcon B, with etafilcon A showing the most change in protein distribution. Albumin removal was highest from etafilcon A >balafilcon A >lotrafilcon B. H2O2 exhibited greater lysozyme removal from etafilcon A compared to both MPS procedures (p0.62). Albumin removal was solely material specific, while all care regimens performed to a similar degree (p>0.69). Conclusions: Protein removal efficiency for the regimens evaluated depended on the lens material and protein type. Overall, lens rubbing with MPS before soaking did not reduce the protein content on the lenses compared to nonrubbed lenses (p=0.89). © 2010 Molecular Vision.

Purpose: To determine albumin adsorption profiles and penetration depth of 3 intraocular lens (IOL) materials over time using confocal laser scanning microscopy (CLSM) and radiolabeling. Setting: Centre for Contact Lens Research, School of Optometry, and Department of Biology, University of Waterloo, Waterloo, Ontario, Canada. Methods: Poly(methyl methacrylate) (PMMA), silicone, and foldable hydrophilic acrylic IOLs were incubated in 0.5 mg/mL bovine serum albumin (BSA) for 1, 7, and 14 days. The BSA was conjugated with lucifer yellow VS to allow identification of the protein location by fluorescent imaging with CLSM. Next, the protein uptake was quantified using 2% 125I-labeled BSA. Results: Confocal laser scanning microscopy showed increasing BSA uptake for silicone and PMMA IOLs after 14 days of incubation (P<.05), with an apparent penetration depth of 8.7 μm ± 1.9 (SD) and 9.2 ± 1.4 μm, respectively. For hydrophilic acrylic IOLs, BSA was detected at a depth of 38 ± 7.4 μm after 1 day, followed by an increase to 192.7 ± 16.2 μm after 14 days. Despite the penetration depth into the hydrophilic acrylic IOLs, quantitative results confirmed that PMMA and hydrophilic acrylic deposited significantly less BSA (mean 278.3 ± 41.7 ng and 296.5 ± 33.1 ng, respectively) than silicone IOLs (mean 392.6 ± 37.6 ng) (P<.05). Conclusions: Silicone and PMMA IOL materials showed BSA sorption near the lens surface only, while BSA penetrated deep into the hydrophilic acrylic IOL matrix. Combining the qualitative CLSM method and quantitative radiolabeling technique provided detailed information on protein interactions with implantable biomaterials. © 2009 ASCRS and ESCRS.

PURPOSE: To investigate the distribution profile of hen egg lysozyme (HEL) through poly-2-hydroxyethyl methacrylate (pHEMA)-based lens materials and silicone hydrogel (SH) lens materials using confocal laser scanning microscopy (CLSM). METHODS: Five silicone SH materials (balafilcon A, lotrafilcon A, lotrafilcon B, galyfilcon A, senofilcon A) and four pHEMA-based materials (alphafilcon A, etafilcon A, omafilcon A, vifilcon A) were incubated in 1.9 mg/ml protein solution for 24 hours. The protein solution consisted of HEL, which was conjugated with either fluorescein isothiocyanate (FITC) or lucifer yellow VS dilithium salt (LY). CLSM (Zeiss LSM 510 META) identified the location of the fluorescently labeled protein by using 1 micro m depth scans through the lens. In a second experiment, lenses were incubated with 2% (125) I labeled HEL to determine the amount of deposited protein on each lens. Both techniques were combined to describe the individual HEL profiles. RESULTS: After the incubation in fluorescently labeled HEL, all pHEMA-based materials and the SH material balafilcon A accumulated protein throughout the entire lens material, while, for the SH lenses lotrafilcon A and lotrafilcon B, HEL was primarily detected on the lens surface alone. Differences in protein uptake pattern due solely to the two conjugated dyes were most apparent for the SH materials galyfilcon A and senofilcon A; HEL was detected throughout these lenses when conjugated with LY but accumulated primarily on the surface when conjugated with FITC. CONCLUSION: CLSM in combination with a radiolabel technique can describe both the location and degree of protein deposition on different contact lens materials.

During contact lens wear, tear film components such as lipids, mucins and proteins tend to deposit on and within the lens material and may cause discomfort, reduced vision and inflammatory reactions. The tear film protein that has attracted most interest when studying contact lens deposition is the small (14 kDa), positively charged protein lysozyme. Albumin, which is a much larger protein (66 kDa) with an overall net negative charge is also of interest, and shows very different adsorption patterns to lysozyme. The concentration of albumin in the tear film is relatively low compared to the concentration in blood serum, but this value increases markedly under various conditions, including when the eye is closed, during contact lens wear and in various dry eye states. Gaining an understanding of the manner in which albumin deposits on biomaterials is of importance for contact lens wear, as well as for other medical applications where HEMA-based materials are used for implants, artificial blood vessels or drug delivery devices. This review paper summarizes the impact of individual material compositions, water content, hydrophobicity and electrostatic attraction on the adsorption behavior of the protein albumin.

Purpose. To develop a novel in vitro method to detect the depth of penetration of the tear film protein albumin into contact lens materials using confocal laser scanning microscopy (CLSM).
Methods. A poly-HEMA-based hydrogel (etafilcon A) and a silicone hydrogel material (lotrafilcon B) were examined. In vitro, bovine serum albumin (BSA) was labeled with 5-(4,6-dichloro-s-triazin-2-ylamino) fluorescein hydrochloride (DTAF). The lenses were incubated in this protein solution (0.5 mg/ml) at 37°C. After 1 and 7 days incubation, the lenses were examined using CLSM (Zeiss 510, config. META 18) and the location of the fluorescently labeled BSA was identified.
Results. BSA adsorption on the surface and penetration into the lens matrix occurred at a higher concentration for etafilcon compared to lotrafilcon (p < 0.001). For both materials, BSA was detected on the surface after 1 day of incubation. Significant levels of BSA were detected within the matrix of etafilcon after as little as 1 day (p < 0.001), but no BSA was detected in the matrix of lotrafilcon at any time (p > 0.05).
Conclusion. CLSM can be successfully used to examine the depth of penetration of fluorescently labeled proteins into various hydrogel polymers. Our results show that etafilcon lenses both adsorb BSA on the surface and absorb BSA within the matrix, whereas lotrafilcon B adsorbs small amounts of BSA on the surface only.

E-abstract 80029

E-abstract 5377

E-Abstract 060092