Lens Solutions Research Articles

Cytotoxic and Inflammatory Effects of Contact Lens Multipurpose Solutions on Human Corneal Epithelial Cells

Multipurpose solutions (MPSs) are the leading method for cleaning and disinfecting soft contact lenses (CLs). During recent years, numerous clinical studies have evaluated the MPS damage to the ocular surface. This study examined the cytotoxic and the inflammatory effects of MPSs and hydrogen peroxide disinfection system (H202) compared to appropriate controls on human corneal epithelial (HCE) cells. Primary cultured HCE cells were exposed to eight different commercially available MPS products (MPS A, ReNu MultiPlus®; MPS B, Opti Free® EverMoist; MPS C, Solo-care Aqua®; MPS-D, Complete®; MPS-E, Unica Sensitive®; MPS-F, Options Multi®; MPS-G, Biotrue®; MPS-H, COMPLETE® RevitaLens). Morphological changes and cytotoxic effects were examined with FITC-Annexin V/PI and MTT assays. The protein contents of the inflammatory cytokines interleukin (IL)-1β, TNF-α, IL-6 and IL-8 were examined by multiplex fluorescent bead immunoassay (FBI), and the mRNA expression was examined by real time PCR. Lipopolysaccharide (LPS) with 500 ng/ml CD14 and 500 ng/ml LBP (LPS complex), polyinosinic: polycytidylic acid (Poly I:C) and un-neutralized H202 served as positive controls, respectively. Phosphate-buffered saline (PBS) was added as a negative control. The study demonstrated that most of the MPSs induced varying degrees of cytotoxicity to HCE cells, and increased production of pro-inflammatory cytokines compared to the negative control. In addition, several MPS increased the mRNA level of inhibitory factor-κBα (1-κBα). Among the various MPSs, MPS-H induced the highest protein contents of the pro-inflammatory cytokines (14.37±2.2-fold for TNF-α, 41.39±2.5-fold for IL-1β and 5.24±0.6-fold for IL-6) compared to the negative control (p<0.05). In contrast, no significant differences were noted between the neutralized H202 and the negative control. We conclude that most of the currently used MPSs induce significant damage and inflammatory response in corneal epithelial cells. MPS-induced inflammation was mediated through NF-κB signal transduction. This study demonstrates for the first time inflammatory responses at the molecular level in primary HCE cells following exposure to a large series of commercially available and commonly used MPSs. These findings strongly suggest that certain MPSs may be partially involved in the pathogenesis of contact lens intolerance. Therefore, we recommend that practitioners advise patients as to the preferable disinfecting contact lens solutions, and to consider using the hydrogen peroxide disinfection systems instead.

Silicone Hydrogel Lens–Solution Interaction and Inflammation

Daily wear of silicone hydrogel (SiHy) lenses using marketed lens care solutions results in varied levels of anterior eye inflammation. There are several pathways that may lead to corneal and conjunctival inflammation that include mechanical, allergic, and pathogen driven stimuli. The hypotheses and evidence for these various models of inflammation will be discussed in the context of SiHy lens designs and materials and lens care solution components.

2. Contact lens care and ocular surface homeostasis

The early focus of contact lens wear and ocular health was on oxygen delivery. However, as we learn more about how the eye works, and investigate how the contact lens interacts with the cornea, the role of the tear film has risen in prominence. A healthy tear film is critical for normal ocular homeostasis, and abnormalities of the tear film are the primary cause of dry eye. In order to improve patient eye health and comfort during lens wear, we need to further elucidate the relationship among contact lenses, contact lens solutions, the tear film, and the corneal epithelium, and find ways to maintain homeostasis of the ocular surface. In this section, we review the latest data and opinions on this complex relationship between contact lenses and lens care solutions.

Effect of contact lens solutions on the antimicrobial efficacy of human tear proteins extracted from contact lenses

Purpose: This group previously reported that proteins recovered from conventional hydrogel contact lenses retain their antimicrobial function, with the extent of this activity depending on the care solution used to treat lenses during wear. This study used tear proteins recovered from PureVision silicone hydrogel lenses to examine how antimicrobial activity is affected by carrying out the assays directly in a disinfecting solution containing Polyaminopropyl Biguanide and Polyquaturnium-1, as well as the individual components of the multipurpose solution (MPS).

A Preservative-and-Fluorescein Interaction Model for Benign Multipurpose Solution–Associated Transient Corneal Hyperfluorescence

Multipurpose contact lens solution (MPS)/preservative-associated transient corneal hyperfluorescence has been suggested to represent corneal injury. To determine the validity of this assumption, the molecular-level interactions of common disinfectants in soft contact lens MPS and the corneal epithelium using an in vitro model were assessed. A liposome-based model of the corneal epithelial surface was developed and used to assess the interactions of polyhexamethylene biguanide (PHMB), polyquaternium-1 (PQ-1), and fluorescein with membrane components and the effects of PHMB and PQ-1 on membrane integrity. The fluorescence anisotropy (a measure of interactions between molecules) was determined. Liposome integrity was assessed by measuring the liposome melting point temperature. Free fluorescein did not associate with the liposome (P>0.4). Both fluorescein-tagged PHMB and PQ-1 associated with liposomes (P<0.002 and P≤0.01, respectively); however, only PHMB induced free fluorescein association with membrane components. At physiological temperature, no significant shift in the melting point temperature was observed when liposomes were exposed to PHMB from 0 to 100 ppm (P>0.05). In contrast, exposure of >7 ppm PQ-1 disrupted the liposomes. Based on this study, PHMB-to-liposome bilayer interaction is nondestructive, even at concentrations 100 times higher than found in commercially available MPS products. In contrast, PQ-1-to-liposome bilayer interaction led to liposome disruption. This study presents molecular-level evidence to support that preservative-associated transient corneal hyperfluorescence is a benign transient phenomenon and its evaluation clinically may be an ambiguous strategy for determining biocompatibility and cell surface integrity.

The clinical performance of three contact lens solutions in symptomatic and asymptomatic contact lens wearers

Purpose: To investigate the performance of a new multipurpose solution based on alexidine and polyquaternium-1 (AMO RevitaLens Ocutec) and two current solutions, based on polyquaternium-1 and Aldox (Alcon Optifree RepleniSH) and hydrogen peroxide (CIBA Vision ClearCare).

The Effects of Contact Lens Materials on a Multipurpose Contact Lens Solution Disinfection Activity Against Staphylococcus aureus

To determine the effect of 8 different lens materials on polyhexamethylene biguanide (PHMB) concentration in multipurpose solution (MPS) levels over time and to determine the effect of lenses on lens solution microbial efficacy over time. Silicone hydrogel lenses and conventional hydrogel lenses were soaked in polypropylene lens cases filled with contact lens MPS containing 1 ppm PHMB for 6, 12, 24, 72, and 168 hours. Cases filled with the same solution without lenses were controls. After each time period, solutions from cases with the 8 types of lenses and controls were assayed for activity against Staphylococcus aureus according to International Organization for Standardization-14729 with modifications. Solutions were analyzed for PHMB concentration at each time point. Some of the different lens materials significantly affected the PHMB concentration (P<0.0001) and the biocidal efficacy. Etafilcon A lenses significantly decreased PHMB levels after only 6 hours of lens soak time. The product lot of MPS used was also significant (P<0.0001). Enfilcon A, senofilcon A, and lotrafilcon B lenses did not significantly decrease PHMB levels. The efficacy of MPS was affected by some lens materials and PHMB concentration. Lens materials differ in their effect on PHMB concentration and the subsequent efficacy of the MPS. Over time, some lens materials can significantly reduce the PHMB concentration and the MPS's microbial activity against S. aureus.

Strategies to Optimize Conditions for Testing Multipurpose Contact Lens Solution Efficacy Against Acanthamoeba

To optimize the growth, culture, and life cycle conditions for testing multipurpose solutions (MPS) against Acanthamoebatrophozoites and cysts to better inform the development of an appropriate test protocol. Two strains of Acanthamoeba castellanii were grown using 2 different methods, bacterized and axenic. Amoebae grown from both methods and from both strains were treated with 4 different MPS as trophozoites and cysts, which were generated using 4 encystment methods. Experiments were run in triplicate with controls. A 5-tube most probable number method was used to enumerate the survivors and to determine the log kills. Statistical analysis was performed using effect screening. There was a marked difference in effectiveness among solutions, which varied with growth conditions (P<0.0001) and encystment method. Growth medium affected survival. In addition, there was a significant difference in cyst survival, which was dependent on encystment method (P=0.0013). The strain used was less of a factor in trophozoite resistance to MPS. Cyst resistance to MPS varied depending on which strain was used, but it was not a significant factor. When designing a contact lens solution efficacy testing protocol for Acanthamoeba, care should be taken to control for variables that may distort results. An appropriate protocol should include growing Acanthamoeba bacterized and allowing them to encyst naturally. By choosing optimized testing conditions, a more realistic efficacy of contact lens solutions can be determined which will result in better and safer products on the market.

Material Properties That Predict Preservative Uptake for Silicone Hydrogel Contact Lenses

To assess material properties that affect preservative uptake by silicone hydrogel lenses. We evaluated the water content (using differential scanning calorimetry), effective pore size (using probe penetration), and preservative uptake (using high-performance liquid chromatography with spectrophotometric detection) of silicone and conventional hydrogel soft contact lenses. Lenses grouped similarly based on freezable water content as they did based on total water content. Evaluation of the effective pore size highlighted potential differences between the surface-treated and non-surface-treated materials. The water content of the lens materials and ionic charge are associated with the degree of preservative uptake. The current grouping system for testing contact lens-solution interactions separates all silicone hydrogels from conventional hydrogel contact lenses. However, not all silicone hydrogel lenses interact similarly with the same contact lens solution. Based upon the results of our research, we propose that the same material characteristics used to group conventional hydrogel lenses, water content and ionic charge, can also be used to predict uptake of hydrophilic preservatives for silicone hydrogel lenses. In addition, the hydrophobicity of silicone hydrogel contact lenses, although not investigated here, is a unique contact lens material property that should be evaluated for the uptake of relatively hydrophobic preservatives and tear components.

Potential Resistant Morphotypes of Acanthamoeba castellanii Expressed in Multipurpose Contact Lens Disinfection Systems

The free-living amoeba Acanthamoeba castellanii is a rare cause of contact lens-associated microbial keratitis. The cyst stage of this amoeba is noted for its resistance to disinfection by multipurpose contact lens solutions (MPS). This report examines and reviews the potential survival modes of A. castellanii in MPS. Trophozoites of A. castellanii (ATCC 30234) at densities from 10 to near 10 were incubated in 3 different MPS in a laminar flow hood for 24 hours at ambient temperatures. The dried films of MPS and phosphate-buffered saline (PBS) controls were examined before and after the addition of a peptone-yeast extract-glucose recovery broth (PYG) for the presence of amoeboid trophozoites and resistance stages over at least 7 days. The parallel exposure of trophozoites to MPS without evaporation or addition of PYG was similarly examined. Amoeboid trophozoites were not recovered in PYG nor were cyst-like structures observed in any MPS with near 10 densities. Progressively with 10 to 10 trophozoites, varied survival modes, particularly aggregates of trophozoites associated with cyst-like structures and occasional amoeboid forms and double-walled cysts with ostioles, became more evident. These morphotypes were most prominent after evaporation and typically first observed in the PYG. Trophozoites of A. castellanii near 10 and progressively to 10 densities are capable of expressing a variety of "short-term" survival modes in MPS, notably with the added stress of evaporation. Expression of these alternate survival modes in MPS may relate, in part, to contamination of contact lens cases and difficulties in developing standardized MPS efficacy tests.

Effect of Multipurpose Solutions on Cell Morphology and Cytokine Production by Corneal Epithelial Cells

We sought to determine whether human corneal limbal epithelial cells (HCLE) differ in their physiological response and production of inflammatory mediators during exposure to two multipurpose disinfecting solutions that differ in only four excipients. HCLE were exposed to dilutions (1%-20%) of OPTI-FREE Express and OPTI-FREE RepleniSH for 2, 6, or 18 h. Cell numbers were measured using CyQuant and metabolic activity assays. Morphology, viability, and apoptotic changes were examined by confocal microscopy after staining with Calcein AM/propidium iodide or Hoechst 33,342/propidiun iodide/YO-PRO-1. Cytokine responses and arachidonic acid metabolites were examined by enzyme-linked immunosorbent assay. OPTI-FREE Express showed greater reductions in corneal cell metabolic activity (up to 3-fold) than OPTI-FREE RepleniSH over 18 h. Cells exposed to OPTI-FREE Express were highly vacuolated, whereas those exposed to OPTI-FREE RepleniSH had morphology consistent with a presumed apoptotic response. OPTI-FREE Express elicited higher levels of interleukin (IL)-6 (maximum 4225 ± 300 pg/mL) and IL-8 (maximum 1094 ± 250 pg/mL) from cells than OPTI-FREE RepleniSH (maximum of 1717 ± 225 pg/mL and 930 ± 300 pg/mL, respectively) at all concentrations tested after 18 h. HCLE did not produce leukotriene B4 or prostaglandin E2 on stimulation, and IL-1β was produced in low levels, but its production was not different between multipurpose disinfecting solution types. The findings reported here are the first to describe the pattern of cytokine production produced in corneal epithelial cells in response to contact lens solutions. Alteration of only four excipients in the formulations had such effects on corneal epithelia; however, these differences do not explain differences in the incidences of corneal infiltrative events between these solutions. This might indicate that changes in the rates of infiltrative events result from interactions of the solutions with the contact lens surface, or directly with the corneal immune system, or as the result of gram-negative contamination of lens cases.

Comparison of the effects of different lens-cleaning solutions on the protein profiles of human conjunctival cells

Many contact lens wearers suffer from dry eye syndrome. Previous studies show significant changes in the protein composition of the tears depending on the lens-cleaning solution used. Therefore, the aim of this study was to detect the influence of different lens solutions on conjunctival cells, cells also involved in tear film composition. Conjunctival epithelium cells (HCjE; IOBA-NHC) were exposed to medium containing Complete® Multipurpose Solution Easy Rub® Formula (Complete) (AMO) or Opti-Free® Express Multipurpose Disinfecting Solution (Opti-Free) (Alcon) in different concentrations (0.1, 0.5, 1, 2.5%) for 12 and 24h. Apoptosis and necrosis using FACS and protein profiles of the cells using SELDI-TOF-MS and MALDI-TOF-MS were measured. Multivariate statistics were calculated to detect the most significant changes. Complex protein profiles were measured with SELDI-TOF-MS and MALDI-TOF-MS. Significant differences of protein profiles between control and treatment cells were detected after 12 and 24h, although cells incubated with Complete showed significantly fewer changes than cells incubated with Opti-Free, also showing concentration-dependent changes of some significantly changed proteins, e.g., protein at 6,736Da (p > 0.002). Cells incubated with Complete showed significantly less apoptosis or necrosis in comparison to control cells (p < 0.05), whereas cells incubated with Opti-Free showed significantly more (p < 0.05). We were able to demonstrate that Complete shows very little effect on the protein profiles of conjunctival cells in comparison to Opti-Free, where the cells showed very large protein profile changes, and apoptosis and necrosis of the cells was increased. These results are in concordance to clinical studies showing that the use of Complete solution made the tear film proteins similar to those of people not wearing contact lenses. Therefore, we believe that Complete is less aggressive and should provoke fewer side-effects, such as dry eye syndrome, for the users.

In vitro biocompatibility assessment of multipurpose contact lens solutions: Effects on human corneal epithelial viability and barrier function

To explore the in vitro effects of multipurpose contact lens solutions (MPSs) on corneal epithelial barrier function and viability. Human corneal epithelial cells (HCEpiC) were exposed to 50% MPSs A-G. Viability was determined using metabolic activity, protease release and caspase assays. Barrier function was evaluated using immunostaining for the tight junction protein zonnula occludens-1 (ZO-1) and resistance measurements. MPS A and G did not affect HCEpiC monolayer viability after 2 h, while MPSs B-F significantly decreased viability. There was a significant decrease in stratified HCEpiC viability after exposure to MPSs B-E for 2 h, while there was no effect of MPS A. After exposure of HCEpiC monolayers to MPS A, F or G for 30 min, ZO-1 staining appeared similar to control. HCEpiC exposed to MPSs B and C demonstrated tight junction breakdown. There was no significant change in HCEpiC monolayer resistance after exposure to MPS A or F for 2 h, while MPSs B-E and G reduced resistance. After exposure to MPS A-E, stratified HCEpiC resistance was significantly decreased after 2 or 4 h. The decrease in resistance was significantly less with MPS A as compared to the other MPSs. MPSs caused varying modifications to cell viability and barrier function in monolayer and stratified HCEpiC. MPS A did not alter monolayer HCEpiC viability or barrier function, while MPSs B-G caused significant decreases of at least one parameter. Furthermore, MPS A had significantly less effect than MPSs B-E on viability and barrier function of stratified HCEpiC.

Antifungal efficacy of soft contact lens disinfecting solutions against Fusarium solani and Candida albicans

Purpose: The aim was to evaluate the disinfection properties of six multi‐purpose contact lens disinfection solutions (MPDS) available in Iran against Fusarium solani and Candida albicans, based on the International Organization for Standardization (ISO) 14729 guidelines.Methods: Three lots each of six multi‐purpose solutions, namely, Renu MultiPlus, Solo Care Aqua, All‐Clean soft, Contact All In One Advanced, Hippia, Ginza Multi‐Purpose Solution and 0.9% normal saline solution were inoculated with standard strains of Fusarium solani (ATCC 36031) and Candida albicans (ATCC 10231). Surviving fungi were determined by counting colony‐forming units on recovery plates at specified times. The mean log reduction in fungal numbers at the manufacturer's minimum recommended disinfection time was determined and compared with the criteria for stand‐alone disinfection products for each MPDS against each fungal strain.Results: Renu MultiPlus, Contact All In One Advanced and All‐Clean Soft were effective in achieving the mean 1.0 log reduction in fungal numbers, based on ISO 14729 stand‐alone primary acceptance criteria. Solo Care Aqua failed to achieve the ISO 14729 stand‐alone primary acceptance criteria for Candida albicans. The other solutions were not effective against test microorganisms after the specified times.Conclusions: There were differences in the rates of disinfection efficacy. Generally, Renu MultiPlus, Contact All In One Advanced and All‐Clean Soft exceeded the primary acceptance criteria of the ISO 14729 guidelines for stand‐alone contact lens solutions.

An Outbreak of Acute Post-Cataract Surgery Pseudomonas sp. Endophthalmitis Caused by Contaminated Hydrophilic Intraocular Lens Solution

To report the investigation for the source of infection and the clinical course and treatment response of 11 cases of acute post-cataract surgery endophthalmitis that developed during an outbreak. Retrospective, consecutive, interventional case series. Eleven patients who developed acute postoperative endophthalmitis after an uneventful cataract surgery with intraocular lens implantation from September 6 to 29, 2010, at a tertiary eye care center in South India. Aqueous aspirates, vitreous aspirates, and environmental surveillance specimens were sampled. All specimens were subjected to smear and culture. Positive cultures were subjected to antibiotic susceptibility. Genotypic diversity was determined by polymerase chain reaction (PCR) with enterobacterial repetitive intergenic consensus (ERIC) primers of each strain and was used to establish the clonal relationship between clinical and environmental isolates. The clinical patterns were analyzed. Positive microbiology, molecular diagnostic similarity among the culture positive endophthalmitis cases, and surveillance specimens. Aqueous and vitreous samples showed gram-negative bacilli in the smears of 8 of 11 eyes, and cultures grew Pseudomonas aeruginosa in 5 of 11 eyes. Among the samples from various surveillance specimens cultured, only the hydrophilic acrylic intraocular lenses and their solution grew P. aeruginosa, with antibiotic susceptibility pattern identical to the clinical isolates. The isolates from the patients and the intraocular lens solution revealed matching patterns similar to an American Type Culture Collection (ATCC) strain of P. aeruginosa on ERIC-PCR. The intraocular lenses of the same make were discontinued at our hospital, and the endophthalmitis did not recur. The final visual acuity improved to ≥ 20/50 in 8 of 11 patients (72.7%). One patient developed retinal detachment, but was treated successfully, and 2 other patients progressed to phthisis bulbi. Positive microbiology and the ERIC-PCR results proved that contamination of hydrophilic intraocular lenses and the preservative solution was the source of infection in this outbreak. Early detection and a planned approach during the outbreak helped us to achieve good visual and anatomic outcomes, even though the offending organism was identified as P. aeruginosa.

Confoscan: An ideal therapeutic Aid and screening tool in acanthamoeba keratitis

Although present worldwide, Acanthamoeba keratitis (AK) is a rare condition. It is a protozoal infection of the eye that is generally caused by wearing contaminated contact lenses or lens solutions. Confoscan and confocal scanning laser tomography (CSLT) are in vivo noninvasive diagnostic tools which provide high definition images of corneal microstructures. Laser in situ keratomileusis (LASIK) is a very common refractive surgery. We report a case series in which the first patient had contact lens induced Acanthamoeba keratitis with corneal epitheliopathy that was unresponsive to conservative treatment. Epithelial debridement was performed based on confoscan findings which confirmed the presence of Acanthamoeba cysts. Subsequently, the cornea re-epithelialized over two days. Another patient had CSLT prior to the LASIK which showed stromal cyst-like structures suggestive of Acanthamoeba keratitis. Four months after medical therapy, repeat CSLT was negative for Acanthamoeba cysts. Third patient was diagnosed with Acanthamoeba infection after undergoing lamellar keratoplasty. CSLT should be used as a screening procedure prior to any corneal refractive surgery to detect and treat protozoal and other infections preoperatively.

A New Method for Evaluation of Compatibility of Contact Lenses and Lens Cases With Contact Lens Disinfecting Solutions

The purpose of this article is to describe new methodology, antimicrobial efficacy endpoint methodology to determine compatibility of contact lens solutions, lens cases and hydrogel lenses for disinfection (AEEMC), to evaluate the effect of a contact lens and a lens case on disinfection efficacy, and to present the ring test used to justify the use of the method in multiple laboratories. A prototype solution containing chlorhexidine as the disinfecting agent and four representative lens types (group I and IV hydrogels and two silicone hydrogels) were used in these ring tests. Five laboratories participated in the chemical and microbiologic analyses. The residual chlorhexidine in lens cases containing the contact lenses was determined using high-performance liquid chromatography; uptake by the lenses was then determined by extrapolation. For the microbiologic part of the study, a contact lens was placed in the well of the lens case, inoculated at 10 to 10 cfu (colony forming units) per lens with microorganisms in 10% organic soil. The microorganisms, Pseudomonas aeruginosa, Serratia marcescens, Staphylococcus aureus, Candida albicans, and Fusarium solani, were prepared as in International Organization for Standardization (ISO) 14729. After a 3- to 10-min exposure time, the prototype solution was dispensed into each well. Aliquots of the inoculated solutions were removed at 4 and 24 hrs and 7 and 30 days and cultured in neutralizing media for determination of survivors; lenses were also cultured for survivors. Chemical uptake data confirmed the differences observed in kill of the challenge organisms according to lens type. It was observed that the culturing of the solution provided adequate data to show the effect of a lens on disinfection efficacy of a lens care product. The findings of the ring test indicated that the separate culturing of the contact lenses is not necessary for routine assessment. The methodology in the November 12, 2008, draft standard (AEEMC), meets the stated objective of demonstrating the effect of a contact lens on the disinfection efficacy of a simulated lens care product. This method, used in combination with the methodology in ISO 14729, should provide for a more robust evaluation of applicable contact lens care disinfecting products.

Contact lens-related microbial keratitis: how have epidemiology and genetics helped us with pathogenesis and prophylaxis

Contact lens wear is a common predisposing factor in microbial keratitis and is one of the two preventable risk factors for corneal infection in a working age population. Our understanding of the prevention and prophylaxis of contact lens-related corneal infection is informed by recent epidemiological studies describing the incidence of and risk factors for the disease, the effect of causative organism on disease severity, and an appreciation of individual immune profiles in susceptibility to and severity of the disease. Although contemporary contact lenses have not reduced the overall incidence of keratitis, a reduction in morbidity may be achievable through recognition of appropriate risk factors in severe disease, including avoiding delays in presenting for appropriate treatment, and attention to storage case hygiene practise. Severe keratitis is most commonly associated with an environmental causative organism, and daily disposable lenses are associated with less severe disease. Pseudomonas aeruginosa remains the commonest cause of contact lens-related corneal infection probably because of its unique virulence characteristics and ability to survive in the contact lens/storage case/ocular environment. In two recent outbreaks of contact lens-related infections, there has been a strong association demonstrated with particular contact lens solutions. Since the recall of these specific contact lens solutions, the rate of Acanthamoeba keratitis has remained above the expected baseline, indicating unidentified risk factors that may include environmental exposures. Individual differences in susceptibility to microbial keratitis may be partly explained by differences in single-nucleotide polymorphisms in certain cytokine genes, particularly those with a proven protective role in corneal infection.

72 A plasma etching and gravimetric monitoring system (PEGMS) for lipid and protein location studies

Purpose: In the tear film lysozyme makes up approximately 35% of the total protein detected and is widely implicated in contact lens spoilation. It functions primarily as a bacteriolytic enzyme in the innate defense of the ocular surfaces. Recently interest in this protein has focused on its ability to function effectively in the presence of contact lens and care solution products. Method: The classic methods based on its interaction with, and the lysis of, the Grampositive bacterial strainMicrococcus lysodeicticus,measured by turbidity at 450nm, were used to investigate lysozyme antimicrobial activity. In addition a fluorescence-based assay which measures the digestion products from a DQTM lysozyme substrate, with excitation maxima at 494nm and fluorescence emission maxima at 518nm, was assessed. Results: Disadvantages of turbidity based assayswere apparent. Additionally many studies use egg white lysozymes as a cheaper assay alternative to human sourced protein, but in addition to compositional variation, human and chicken lysozymes differ in enzymatic activity with a 4:1 ratio respectively. Batch to batch variation in lysozyme preparations was also observed. The kinetic fluorescencebased assaywhich was able to detect activity down to 20U/ml was established and was used to assess the effects that extraction techniques and sample preparation have on the outcome of the activity data. Conclusions: An investigation into the true nature and extent of lysozyme denaturation, which is essential to determine the importance of this phenomenon and its impact on tear film anti-microbial function, was facilitated by more sensitive and varied analytical techniques.

Effect of contact lens solutions on the structural and functional integrity of lens bound tear proteins

Effect of contact lens solutions on the structural and functional integrity of lens bound tear proteins