Alan Ng | University of Waterloo (original) (raw)

Papers by Alan Ng

Research paper thumbnail of Impact of in vitro Tear Film Composition on Lysozyme Deposition and Denaturation

Purpose To study the impact of lactoferrin and lipids on the kinetic deposition and denaturation ... more Purpose To study the impact of lactoferrin and lipids on the kinetic deposition and denaturation of lysozyme on contact lens materials. Methods The contact lenses investigated in this thesis included 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]. All lenses were incubated in four solutions: a complex artificial tear solution (ATS); an ATS without lactoferrin; an ATS without lipids; and an ATS without lactoferrin and lipids. Following various time points, all lenses were prepared for lysozyme analysis using the methods below: • To quantify the kinetic uptake of lysozyme to different contact lens materials, I125-radiolabelled lysozyme was added to each incubation solution. Total lysozyme deposition was quantified using a gamma counter. • To study the activity of lysozyme deposited to contact lenses, a fluorescence-based lysozyme activity assay was compar...

Research paper thumbnail of Lysozyme activity on contact lenses and the impact of denatured lysozyme on human corneal epithelial cells

Contact Lens and Anterior Eye

Research paper thumbnail of Case report: Using scleral lens wear to managing severe ocular dryness secondary to an autoimmune disorder

Contact Lens and Anterior Eye

Research paper thumbnail of Surface versus bulk activity of lysozyme deposited on hydrogel contact lens materials in vitro

Contact lens & anterior eye : the journal of the British Contact Lens Association, Jan 30, 2018

To determine and compare the levels of surface versus bulk active lysozyme deposited on several c... more To determine and compare the levels of surface versus bulk active lysozyme deposited on several commercially available hydrogel contact lens materials. Hydrogel contact lens materials [polymacon, omafilcon A, nelfilcon A, nesofilcon A, ocufilcon and etafilcon A with polyvinylpyrrolidone (PVP)] were incubated in an artificial tear solution for 16 h. Total activity was determined using a standard turbidity assay. The surface activity of the deposited lysozyme was determined using a modified turbidity assay. The amount of active lysozyme present within the bulk of the lens material was calculated by determining the difference between the total and surface active lysozyme. The etafilcon A materials showed the highest amount of total lysozyme activity (519 ± 8 μg/lens, average of Moist and Define), followed by the ocufilcon material (200 ± 5 μg/lens) and these two were significantly different from each other (p < 0.05). The amount of surface active lysozyme on etafilcon and ocufilcon ...

Research paper thumbnail of Impact of Tear Film Components on Lysozyme Deposition to Contact Lenses

Optometry and Vision Science, 2012

To investigate the impact of lactoferrin and lipids on the kinetic deposition of lysozyme on sili... more 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). 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 &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp; 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. 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 &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt; 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 &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt; 0.001). Etafilcon A lenses deposited the most lysozyme in all four solutions compared with the rest of the lens types (p &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt; 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 &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt; 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 &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt; 0.001). 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.

Research paper thumbnail of Impact of Tear Film Components on Lysozyme Deposition to Contact Lenses

Purpose. To investigate the impact of lactoferrin and lipids on the kinetic deposition of lysozym... more 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 g) than when incubated in solutions not containing lactoferrin and lipids (more than 11.8 g) (p 0.001). Lysozyme uptake to senofilcon A lenses was higher in ATS w/o Lip (5.3 0.1 g) compared with other solutions (less than 3.9 g) (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 g) compared with the other solutions (more than 642.6 g) (p 0.001). Omafilcon A lenses in ATS w/o Lac accumulated significantly less lysozyme (12.8 1.0 g) compared with the other solutions (more than 14.2 g) (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. (Optom Vis Sci 2012;89:392–400)

Research paper thumbnail of Impact of tear film components on the conformational state of lysozyme deposited on contact lenses

Journal of Biomedical Materials Research Part B: Applied Biomaterials, 2013

To investigate the impact of lactoferrin and lipids on the kinetic denaturation of lysozyme depos... more To investigate the impact of lactoferrin and lipids on the kinetic denaturation of lysozyme deposited on silicone and conventional hydrogel lenses, using a complex artificial tear solution (ATS). 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 incubated in four solutions: an ATS, ATS without lactoferrin, ATS without lipids, and ATS without lactoferrin and lipids. At various time points over a 28-day period, the percentage of active lysozyme per lens was determined using a fluorescence activity assay and an ELISA. After 28 days, the percentage of active lysozyme extracted from etafilcon A lenses in all solutions was significantly higher than all other lens materials (p &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt; 0.001). For lotrafilcon B, senofilcon A, and omafilcon A lenses, lysozyme denaturation was greatest during the first week of incubation and before reaching a plateau (p &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;gt; 0.05). The inclusion of lipids in the ATS significantly increased the lysozyme denaturation on both silicone hydrogel materials (p &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt; 0.001), while in the presence of lactoferrin, lysozyme activity on senofilcon A lenses was significantly higher (p &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt; 0.001). Lysozyme activity on both conventional lenses was not significantly affected by either lactoferrin or lipids (p &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;gt; 0.05). Lactoferrin and lipids have an impact on the denaturation of lysozyme deposited onto silicone hydrogel contact lenses, while conventional hydrogel lenses were unaffected. Future in vitro studies should consider the impact of tear film components when investigating protein deposition and denaturation on contact lenses.

Research paper thumbnail of Optimization of a Fluorescence-based Lysozyme Activity Assay for Contact Lens Studies

Current Eye Research, 2013

Purpose: To optimize a fluorescence-based lysozyme activity assay to investigate the conformation... more Purpose: To optimize a fluorescence-based lysozyme activity assay to investigate the conformational state of lysozyme in solution and to determine the impact of extraction and evaporation procedures and the possible interference of contact lens materials on lysozyme activity. Methods: The fluorescence-based lysozyme activity assay, Enzchek (Molecular Probes Inc, Eugene, OR) which utilizes fluorescently quenched Micrococcus lysodeikticus, was compared to the gold standard, classical lysozyme turbidity assay, using four differently concentrated lysozyme samples (20, 10, 5.0 and 2.0 ng/mL). Furthermore, six differently concentrated lysozyme samples (2.0, 1.0, 0.5, 0.25, 0.125 and 0.01 mg/mL) were quantified using the fluorescence-based assay in the presence of extraction solvents consisting of 0.2% and 0.02% trifluroacetic acid/acetonitrile and following evaporation procedures. Results: A standard curve was generated by the fluorescence-based assay ranging from 2 to 150 ng. The total active lysozyme quantified in the four lysozyme samples was not significantly different between the two assays (p40.05) and the concordance correlation coefficient was determined to be 0.995. However an average discrepancy between the two assays was found to be 0.474 ng, with the turbidity assay typically reporting higher active lysozyme measurements. The sensitivity of the fluorescence-based assay was higher than the classical turbidity assay when quantifying 20 ng or less active lysozyme. Following the extraction and evaporation procedures and the addition of lens extracts, the total active lysozyme recovered was 95% or greater. Conclusions: In comparison to the classical turbidity assay, the fluorescence-based assay is a very sensitive method, making it a favorable technique, particularly when studying contact lens materials that deposit relatively low levels of lysozyme.

Research paper thumbnail of Impact of Tear Film Components on Lysozyme Deposition to Contact Lenses

Optometry and Vision Science, 2012

To investigate the impact of lactoferrin and lipids on the kinetic deposition of lysozyme on sili... more 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). 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 &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp; 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. 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 &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt; 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 &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt; 0.001). Etafilcon A lenses deposited the most lysozyme in all four solutions compared with the rest of the lens types (p &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt; 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 &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt; 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 &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt; 0.001). 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.

Research paper thumbnail of Impact of in vitro Tear Film Composition on Lysozyme Deposition and Denaturation

Purpose To study the impact of lactoferrin and lipids on the kinetic deposition and denaturation ... more Purpose To study the impact of lactoferrin and lipids on the kinetic deposition and denaturation of lysozyme on contact lens materials. Methods The contact lenses investigated in this thesis included 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]. All lenses were incubated in four solutions: a complex artificial tear solution (ATS); an ATS without lactoferrin; an ATS without lipids; and an ATS without lactoferrin and lipids. Following various time points, all lenses were prepared for lysozyme analysis using the methods below: • To quantify the kinetic uptake of lysozyme to different contact lens materials, I125-radiolabelled lysozyme was added to each incubation solution. Total lysozyme deposition was quantified using a gamma counter. • To study the activity of lysozyme deposited to contact lenses, a fluorescence-based lysozyme activity assay was compar...

Research paper thumbnail of Lysozyme activity on contact lenses and the impact of denatured lysozyme on human corneal epithelial cells

Contact Lens and Anterior Eye

Research paper thumbnail of Case report: Using scleral lens wear to managing severe ocular dryness secondary to an autoimmune disorder

Contact Lens and Anterior Eye

Research paper thumbnail of Surface versus bulk activity of lysozyme deposited on hydrogel contact lens materials in vitro

Contact lens & anterior eye : the journal of the British Contact Lens Association, Jan 30, 2018

To determine and compare the levels of surface versus bulk active lysozyme deposited on several c... more To determine and compare the levels of surface versus bulk active lysozyme deposited on several commercially available hydrogel contact lens materials. Hydrogel contact lens materials [polymacon, omafilcon A, nelfilcon A, nesofilcon A, ocufilcon and etafilcon A with polyvinylpyrrolidone (PVP)] were incubated in an artificial tear solution for 16 h. Total activity was determined using a standard turbidity assay. The surface activity of the deposited lysozyme was determined using a modified turbidity assay. The amount of active lysozyme present within the bulk of the lens material was calculated by determining the difference between the total and surface active lysozyme. The etafilcon A materials showed the highest amount of total lysozyme activity (519 ± 8 μg/lens, average of Moist and Define), followed by the ocufilcon material (200 ± 5 μg/lens) and these two were significantly different from each other (p < 0.05). The amount of surface active lysozyme on etafilcon and ocufilcon ...

Research paper thumbnail of Impact of Tear Film Components on Lysozyme Deposition to Contact Lenses

Optometry and Vision Science, 2012

To investigate the impact of lactoferrin and lipids on the kinetic deposition of lysozyme on sili... more 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). 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 &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp; 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. 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 &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt; 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 &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt; 0.001). Etafilcon A lenses deposited the most lysozyme in all four solutions compared with the rest of the lens types (p &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt; 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 &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt; 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 &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt; 0.001). 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.

Research paper thumbnail of Impact of Tear Film Components on Lysozyme Deposition to Contact Lenses

Purpose. To investigate the impact of lactoferrin and lipids on the kinetic deposition of lysozym... more 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 g) than when incubated in solutions not containing lactoferrin and lipids (more than 11.8 g) (p 0.001). Lysozyme uptake to senofilcon A lenses was higher in ATS w/o Lip (5.3 0.1 g) compared with other solutions (less than 3.9 g) (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 g) compared with the other solutions (more than 642.6 g) (p 0.001). Omafilcon A lenses in ATS w/o Lac accumulated significantly less lysozyme (12.8 1.0 g) compared with the other solutions (more than 14.2 g) (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. (Optom Vis Sci 2012;89:392–400)

Research paper thumbnail of Impact of tear film components on the conformational state of lysozyme deposited on contact lenses

Journal of Biomedical Materials Research Part B: Applied Biomaterials, 2013

To investigate the impact of lactoferrin and lipids on the kinetic denaturation of lysozyme depos... more To investigate the impact of lactoferrin and lipids on the kinetic denaturation of lysozyme deposited on silicone and conventional hydrogel lenses, using a complex artificial tear solution (ATS). 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 incubated in four solutions: an ATS, ATS without lactoferrin, ATS without lipids, and ATS without lactoferrin and lipids. At various time points over a 28-day period, the percentage of active lysozyme per lens was determined using a fluorescence activity assay and an ELISA. After 28 days, the percentage of active lysozyme extracted from etafilcon A lenses in all solutions was significantly higher than all other lens materials (p &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt; 0.001). For lotrafilcon B, senofilcon A, and omafilcon A lenses, lysozyme denaturation was greatest during the first week of incubation and before reaching a plateau (p &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;gt; 0.05). The inclusion of lipids in the ATS significantly increased the lysozyme denaturation on both silicone hydrogel materials (p &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt; 0.001), while in the presence of lactoferrin, lysozyme activity on senofilcon A lenses was significantly higher (p &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt; 0.001). Lysozyme activity on both conventional lenses was not significantly affected by either lactoferrin or lipids (p &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;gt; 0.05). Lactoferrin and lipids have an impact on the denaturation of lysozyme deposited onto silicone hydrogel contact lenses, while conventional hydrogel lenses were unaffected. Future in vitro studies should consider the impact of tear film components when investigating protein deposition and denaturation on contact lenses.

Research paper thumbnail of Optimization of a Fluorescence-based Lysozyme Activity Assay for Contact Lens Studies

Current Eye Research, 2013

Purpose: To optimize a fluorescence-based lysozyme activity assay to investigate the conformation... more Purpose: To optimize a fluorescence-based lysozyme activity assay to investigate the conformational state of lysozyme in solution and to determine the impact of extraction and evaporation procedures and the possible interference of contact lens materials on lysozyme activity. Methods: The fluorescence-based lysozyme activity assay, Enzchek (Molecular Probes Inc, Eugene, OR) which utilizes fluorescently quenched Micrococcus lysodeikticus, was compared to the gold standard, classical lysozyme turbidity assay, using four differently concentrated lysozyme samples (20, 10, 5.0 and 2.0 ng/mL). Furthermore, six differently concentrated lysozyme samples (2.0, 1.0, 0.5, 0.25, 0.125 and 0.01 mg/mL) were quantified using the fluorescence-based assay in the presence of extraction solvents consisting of 0.2% and 0.02% trifluroacetic acid/acetonitrile and following evaporation procedures. Results: A standard curve was generated by the fluorescence-based assay ranging from 2 to 150 ng. The total active lysozyme quantified in the four lysozyme samples was not significantly different between the two assays (p40.05) and the concordance correlation coefficient was determined to be 0.995. However an average discrepancy between the two assays was found to be 0.474 ng, with the turbidity assay typically reporting higher active lysozyme measurements. The sensitivity of the fluorescence-based assay was higher than the classical turbidity assay when quantifying 20 ng or less active lysozyme. Following the extraction and evaporation procedures and the addition of lens extracts, the total active lysozyme recovered was 95% or greater. Conclusions: In comparison to the classical turbidity assay, the fluorescence-based assay is a very sensitive method, making it a favorable technique, particularly when studying contact lens materials that deposit relatively low levels of lysozyme.

Research paper thumbnail of Impact of Tear Film Components on Lysozyme Deposition to Contact Lenses

Optometry and Vision Science, 2012

To investigate the impact of lactoferrin and lipids on the kinetic deposition of lysozyme on sili... more 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). 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 &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp; 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. 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 &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt; 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 &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt; 0.001). Etafilcon A lenses deposited the most lysozyme in all four solutions compared with the rest of the lens types (p &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt; 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 &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt; 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 &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt; 0.001). 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.