EARBUDDY Novel PEGylated Tympanostomy Tube for

Biofilm Inhibition

Lars Gustafson1, Byron Ho2, Adit Kothari1, Arun Pingali1, Minhao Zhou2

BioE C1171 & MechE C1172 Structural Aspects of Biomaterials, Spring 2016 University of California, Berkeley

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Executive Summary

Otitis media describes a set of diseases that cause inflammation of the middle ear. Recurrent episodes and severe cases may require the surgical insertion of tympanostomy tubes to correct the problem. Unfortunately, current tympanostomy tube technology has several major drawbacks as well as remission in up to a quarter of the patients. The reason for the recurrence of otitis media is due to the formation of a antibiotic resistant bacterial biofilms well as granular tissue formation which can obstruct the tubes. To prevent both the formation of granular tissue and biofilm, we will coat the surface of the tubes with polyethylene glycol (PEG) to stop protein adsorption, which in turn prevents both buildups. By preventing further episodes of otitis media as well as otorrhea (discharge) our device will reduce costs and surgeries while improving the health of our patients. Of course, safety is important as well, so our device will be built using polytetrafluoroethylene (flouroplastic), which is a strong, biocompatible, and inert material. For FDA approval, the device will need to prove sterilization (performed using ethylene oxide gas), undergo in vitro testing, and show efficacy, which would be a decrease in post-operative otorrhea relative to current tubes via the use of the PEG coating to prevent tissue and biofilm formation. The combination of this flouroplastic material and the subsequent PEG coating is a novel idea and thus not covered under current intellectual property claims. While there are current devices that use PEG as well as devices built with similar materials and design, the unique usage of PEG and the combination of this flouroplastic material with the subsequent PEG coating is a novel idea and thus not covered under current intellectual property claims. We have devised an original, improved Class II tympanostomy tube device and are planning on pursuing FDA approval in the near future.

Introduction and Background

Otitis media refers collectively to several inflammatory diseases that affect the middle ear, with the two most prominent being acute otitis media (AOM) and otitis media with effusion (OME), both of which can be associated with hearing loss. AOM usually causes visible symptoms as well as pain, and is prevalent in small children, while OME typically does not present any symptoms. Typically antibiotics are only recommended in very small children and those with severe disease because the antibiotics often have little effect but do cause side effects. A solution for those with frequent ear disease is the use of tympanostomy tubes,

which are small tubes placed in the eardrum to allow airflow to the middle ear while simultaneously preventing fluid from building up. Tympanostomy tubes are implanted using surgery under general anesthesia so that the patient cannot inadvertently move and impede the procedure. These tubes, otherwise known as grommets, are most commonly made from plastics such as silicon or Teflon and remain in the eardrum for around 2 years or until they spontaneously fall out. One common and major complication caused by the use of grommets is the onset of otorrhea, which is fluid leak/discharge from the ear, which is an issue modern grommet technology is trying to solve.

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Novelty As previously mentioned, our device intends to meet unresolved problems found in current tympanostomy tube devices, namely post-operative otitis media and granulation tissue formation. Our design’s novelty is centered around implementation of a polyethylene glycol (PEG) coating to create an anti-fouling surface on the tube. Although our geometrical and structural designs will be based upon previous tympanostomy tubes, the integration of a PEG coating (PEGylation) allows our design to differ from any existing tympanostomy tube.

Bacterial biofilm inhibition Otitis media, as mentioned in the introduction, is an inflammatory infection in the middle ear. Otitis media in post-operative tympanostomy patients is reported to reoccur at high rates - 23% from Gonzalez et al1. Additionally, post-tympanostomy tube otorrhea - drainage exiting the ear - caused by otitis media has been seen in up to 74% of patients2. This rampant infection rate following tympanostomy tube insertion h ighl ights a severe lack in device effectiveness. Evidence behind otitis media in tympanostomy tube patients has been traced to bacterial biofilm formation3. Once bacteria are allowed to aggregate and form a biofilm, they are no longer treatable by antibiotics. Since most current treatment against otitis media is with antibiotic ear drops, there is clearly a lack in adequate protection against numerous ha rmfu l mic roorgan i sms . Additionally, the continual use of antibiotics may lead to harmful bacterial resistance. Our device intends to directly address the issue of bacterial biofilm formation. As mentioned, this will be achieved through implementation of a PEG coating to the surface of the tympanostomy tubes. Although

there isn’t complete understanding of the mechanism of biofilm formation, it is accepted that the initial step is unspecific protein adsorption. These proteins then lower the energy barrier of bacterial attachment to the surface. Once bacteria attach to the tympanostomy tube surface, they proliferate and when they sense a bacterial presence at a sufficiently high concentration, they begin expressing traits associated with biofilm formation. So if this initial protein adsorption is inhibited, it should also inhibit biofilm formation. PEG has previously been shown to have desirable properties to resist protein adsorption4,5,6. If correctly grafted onto a surface, PEG inhibits protein binding through entropic repulsion and osmotic pressure. This unfavorable entropy is associated with the compressing of chains between the surfaces and the repulsion between the chains on the same surface and the opposing surface. Additionally, if compressed by a protein, the bound water to the ethylene glycol subunit will become unbound. Since each ethylene glycol subunit binds to 2.3 H2O molecules, this creates a highly unfavorable osmotic pressure as the H2O molecules will attempt to re-bind to PEG and force out the protein. Grafting PEG onto the surface of the tympanostomy tube can be achieved by using two solvents with different intramolecular expansion factors (α) . A solvent with α > 1 is a “good” solvent meaning that the solvent-polymer interactions are thermodynamically favorable while a “bad” solvent is characterized by α < 1 meaning that the s o l v e n t - p o l y m e r i n t e r a c t i o n s a r e t h e r m o d y n a m i c a l l y u n f a v o r a b l e . Functionalizing the surface while in a bad solvent will cause the PEG chains to have a higher pinning density as they will not interact with the solvent as well and will not be swollen. This higher pinning density can

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then be placed in a good solvent and it will swell the PEG, create a highly dense layer that will optimally inhibit protein adsorption.

Granulation tissue inhibition Another key issue that our device addresses is granulation tissue formation around the tympanostomy tubes. Granulation tissue is mainly composed of extracellular matrix created by fibroblasts and small blood vessels that forms as a part of the wound healing process. In approximately 5% of post-tympanostomy patients, granulation tissue formation causes obstruction of the tubes1. If complete obstruction occurs, there are cases where surgery is needed to remove the unwanted tissue. The PEG coating can similarly act to inhibit granulation tissue formation. Fibroblastic tissue secretes extracellular matrix into extracellular space to for the granulation tissue7. In order for this extracellular matrix composed of activated fibroblasts and collagen to settle, unspecific bound proteins must first bind to this surface. Because of this necessary step, inhibition of protein adsorption by PEG will also inhibit undesirable tube obstruction by granulation tissue formation.

Tympanostomy Tube Material T h e A r m s t r o n g G r o m m e t Tympanostomy Tube geometry is one of the m o s t c o m m o n l y u s e d a n d w i d e l y recommended tube by doctors8. Our modifications to the tympanostomy tube will therefore be improvements on this design. This design incorporates the use of the material known as Fluoroplastic, or Polytetrafluoroethylene. The rigidity and smoothness of this material aids the ease of the surgical procedure required to implant the device. Added benefits include the material

being biocompatible and inert. This material is also used for its certain non-adhesive characteristics, which are intended to be used to prevent biofilm formation9. Although this material does prevent biofilm formation as attempted by Staphylococcus Aureus (S. Aureus), it fails to prevent biofilm formation due to Pseudomonas Aeruginosa (P. A e r u g i n o s a ) 1 0 . A d d i t i o n a l l y t h e tympanostomy tube is most commonly affected by Streptococcus Pneumoniae (S. Pneumoniae)11. To prevent biofilm formation, as caused by S. Pneumoniae and P. Aeruginosa, we will modify our tube by coating it with a PEG layer.

Figure 1: Marketing picture and slogan for “Earbuddy"

Figure 1 shows our marketing scheme, highlighting the tube’s antibacterial properties and its innocuousness, as a large portion of customers will be children.

Tests & Outcomes For FDA Approval In order for this device to be approved in the 510(k) submission process, there are a few necessary tests needed to be performed. Additionally, the FDA provides specific guidance and tests for antimicrobial tympanostomy tube testing which we will need to perform.

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Sterilization The sterilization method for our tympanostomy tubes will be through ethylene oxide (EtO) gas in a highly controlled environment. This is the most commonly used sterilization method for both antimicrobial and normal tympanostomy tubes. PEG coatings have previously been shown to be resistant to most standard sterilization techniques including EtO gas treatment12-13. This finding supports our desired use of EtO gas.

In Vitro Testing In vitro test data characterizing the spectrum and degree of activity of the PEG coating against cl inical ly important microorganisms is necessary. These important m i c r o o rg a n i s m s a r e S t r e p t o c o c c u s pneumoniae, Haemophilus influenzae, Branhamella catarrhalis, Staphylococcus aureus, Staphylococcus epidermidis, Diplococcus pneumoniae, and Pseudomonas aeruginosa. All of these bacteria have been shown to form biofilms on tympanostomy tubes. In order to simulate bacterial c o n t a m i n a t i o n , m e d i a c o n t a i n i n g physiological concentrations of these microorganisms along with proteins found in the middle ear space was created. Our PEGylated tympanostomy tubes as well as a control tympanostomy tube were then submerged in this solution and removed at bi-weekly time intervals. Contact angle measurements were performed to probe for potential changes in the hydrophobicity in the surface which could correlate to protein adsorption. The surface was then imaged using SEM in order to visualize the potential biofilms formed in regions where the contact angle had changed. We expect to find significantly less biofilm formation with our

device compared to the non PEGylated tube. However, we do expect some increased biofilm formation with longer incubation periods. Our results would show the bacteriostatic properties of the tube.

Decrease in Post-Operative Otorrhea compared to Conventional Tympanostomy Tube For antimicrobial tympanostomy tubes, the FDA requires data showing a decrease in post-operative otorrhea compared to the otorrhea rate in conventional tympanostomy tubes. In order to compare the two tubes, a clinical trial must be performed with some patients implanted with our PEGylated tube and others implanted with the conventional tube. Otorrhea rates will be measured weekly for up to 40 weeks for both tubes. We expect to find the conventional tympanostomy tube to have a maximum post-operative otorrhea rate of approximately 25% while our PEG-ylated tube will have a maximum post-operative otorrhea rate of approximately 6%. This maximum otorrhea rate is comparable to antimicrobial tympanostomy tubes seen on the market14.

Biocompatibility & Granulation Tissue Formation Our tympanostomy tubes would be implanted in the middle ear of animals and evaluated in a 12 month study. As the tubes have already been shown to inhibit biofilm formation, this study will show that there are no significant adverse effects to the surrounding tissue and that there is limited granulation tissue formation. Following the 12 month study, histological samples can be extracted to test for inflammation and a fibrous capsule in order to determine biocompatibility. Analysis of granulation tissue formation can initially be screened

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under a standard microscope then extraction and staining for fibroblasts and collagen deposits would be performed. Though these tests, the biocompatibility and inhibition of granulation tissue formation characteristics of the tubes will be apparent.

Class II FDA Regulatory Process Tympanostomy tubes are considered to be Class II devices according to the Department of Health and Human Services with the Code of Federal Regulations number 874.388015. Because this medical device is deemed to possess minimal risk, it requires a Premarket Notification 510(k) as a part of its regulatory process. The first step in preparing a medical device for manufacturing in the United States is to gain the Food and Drug Administration's (FDA) approval. According to the FDA's overview of the process, the manufacturers and distributors of the device must first perform an Establishment Registration online as part of the Code of Federal Regulations Title 21 which registers any parties involved

in the production and preparation of the medical device. Once the medical device has been approved by the FDA to be marketed, the registration must be renewed annually in order to continue any form of manufacture. Our tympanostomy tube design is based off of an existing Armstrong Grommet model which was previously placed categorized in the Class II medical device. Therefore, our design will pursue a Premarket Notification 510(k) from the FDA as opposed to a Premarket Approval which are usually performed for Class III devices. According to the FDA website, a 510(k) is required for our design since we are pursuing "a change or modification of a legally marketed device and that change could significantly affect its safety or effectiveness"16. The PEG coating that we are implementing is intended to reduce biofilm formation and to prevent bacterial binding to the tympanostomy tube, but it has yet to be applied for this purpose yet. Therefore, our usage of the PET coating is considered a new modification of an already marketed device.

Figure 2: FDA approval flow chart

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Our group falls under the category as "specification developers introducing a device to the U.S. market"17. We are engineers who are creating a modified design of an existing product and who are not responsible for the manufacturing portion of the design. Therefore, we will submit our 510(k) under this clause. Certain Class II devices do not require a 510(k) submission, so we must verify that the device is not included in the 510(k) Exempt list. If tympanostomy tubes are found to be in this group, then a 510(k) submission would not be necessary. Tympanostomy tubes possess the CFR number 874.3880. Under the Medical Device Exemptions 510(k) list Part 874 which include ear, nose, and throat devices, our device group is not listed, indicating that we must submit a 510(k) in order to market our design. Prior to submitting an official Premarket Notification 510(k) to the FDA, we must request a letter of substantial equivalence from the FDA. This letter acts as an order for the 510(k) and verifies that the device is substantially equivalent to a similar device that has been deemed safe and has already been successfully marketed in the United States18. Once this order has been received, we will be ready to submit a 510(k) application. Substantially equivalence is a demonstration that "a new device is at least as safe and effective as the predicate"19. According to the FDA requirements, our device can be considered substantially equivalent to another legally marketed device under the conditions that it "(1) has the same intended use as the predicate and (2) p o s s e s s e s d i f f e r e n t t e c h n o l o g i c a l characteristics and the information submitted to FDA"20.

The FDA provides a list of twelve organizations with Accredited Persons who can perform a third party review of our device. These services are reserved for Class I and Class II devices. However, the tympanostomy tubes (874.3880) are not listed under the consultable devices so we will not use this resource. After the 510(k) has been submitted, there is a thirty day period in which the FDA must respond. If the device is approved, then it can be found in the FDA's monthly listings of cleared devices. At this point, the device is cleared to be marketed. Manufacturers must be sure to follow Quality System regulations in order to follow Good Manufacturing practices. Following the proper labeling regulations is also necessary to avoid legal troubles. If there have been any incidents involving death or serious injury in which the medical device might be a cause of, the FDA must be informed under the Medical Device Reporting program. There are still many procedures to adhere to even when the FDA has cleared our device for marketing and production.

Figure 3: Patent style graphic indicating novel features

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Intellectual Property Our patent will revolve around the novel use of a PEG coating for tympanostomy tubes, labelled as Feature 4 in Figure 3. As previously explained in more detail, the PEG coating will prevent biofilm formation on the device and therefore lower otitis media recurrence rates. There are numerous patents related to our modified design. A few of these features will be incorporated into our design while other similar features will be explained to not be of concern for infringement.

Antimicrobial The antimicrobial tympanostomy tube is seemingly the most related to our modified design. The goal of this device is to prevent any bacteriostatic infections that may occur post-operation. The main player in the prevention of infection is the use of a silver compound that coats the side walls of the tube. The tube walls have a high gas permeability rate and are made of highly flexible thermoplastic or a thermoset resin. The silver compound would be 0.5 to 15% of the tube by weight. This is a traditional method used to prevent a buildup of bacteria and defend against long term infections. For our modified device, copyright infringement won’t be an issue. Even though the end goal is the same, preventing bacteriostatic infection from biofilm fo rmat ion , our dev ice wi l l no t be incorporating the traditional method of using a silver compound to promote antimicrobial activity. Instead, the surface of our tube will coated with PEG as explained previously.21

PEG Detlef et al created a tympanostomy tube that incorporates PEG in its design, which is the only other tympanostomy tube patent using this polymer. This design has

flanges that include recesses to hold pharmaceutical products. These flanges are designed to be resorbable and eventually follow through and be resorbed. PEG is used as a copolymer alongside starch acetate for the flanges. PEG is incorporated due to its own resorbable properties and it plays no role as a bacteriostatic. For our modified device, copyright infringement won’t be an issue. Although both our device and this device incorporate the use of PEG, the intention and outcome are completely different. This device uses PEG for its resorbable properties and intends for it to eventually be resorbed into the body. It also is built into the device material as a copolymer, instead of a coating which is how we are intending to use it. We will use the coating to prevent biofilm formation due to the PEG’s anti-fouling properties. We are not using the PEG for its resorbable properties, nor do we want it to be resorbed22.

Armstrong Design The tympanostomy tube design that has been recommended by doctors to use is the Armstrong Grommet Tympanostomy tube8. This patent contains specific geometry to show how the device is designed and placed within the ear. The geometries include the outer flange (Feature 3), inner flange (Feature 1), and the mid-tube (Feature 2)23. Dimensions of the tube were also obtained to build a model of the tube24. The material used in certain designs is fluoroplastic, due to its rigid and non-adhesive properties23. This design has been recommended and commonly used without significant issue in terms of mechanical design failure, and therefore will be used in our modified device. Because it has worked so well, we intend to license the patent and use it as the mechanical design upon which we will make our modifications8.

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Surgical Procedure The surgical procedure undergone to implant the tympanostomy tube is the most commonly done surgery in the United States25. This procedure, known as Tympanostomy, requires that the patient be placed under anesthetics. Once the patient is completely under anesthetics, an incision is made in the patient’s affected ear drum. The incision made in the eardrum, more specifically the tympanic membrane, is then widened into a hole that is large enough to tightly fit the diameter of the tympanostomy tube. The tube is then inserted into this hole and allowed to set such that part of the tube is within the eardrum while the other part is outside of it. The hole within the tube allows air and fluids to pass through the eardrum as a response to build up of any sort. A separate surgical procedure is required to replace a tympanostomy tube if it has fallen out or to remove a tympanostomy tube. Because this procedure is quite simple and has proven to be very effective in implanting the device, we intend to license the patent of this procedure to avoid litigation, while still allowing our modified device to aid patients26.

Conclusion Our novel PEGylated tympanostomy tube allows for inhibition of biofilm formation during the lifetime of the implant, lowering otitis media infection rate in post-operative patients. Our PEG coating inhibits protein binding through entropic repulsion and osmotic pressure. Based on our in vitro testing and comparison with conventional tympanostomy tube, the anti-fouling characteristics of our coating shows a significant improvement regarding biofilm formation and eventual infection. In order to bring our device to market, we must follow a

specific FDA pathway. This pathway begins with registration with FDA followed by filing for a 510(k). Tests for the 510(k) submission were previously outlined. Since our device has significant geometric and functionality similarities and to previous devices, substantial equivalence can be claimed. One these steps are approved, marketing and production can begin. Our novelty stems from using a PEG coating as a biofilm inhibition surface. Some features, such as the Armstrong device geometry and a typical tympanostomy tube surgical procedure are already patented and we will need to license these from the respective patent holders.

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References

1. Gonzalez, C., Arnold, J.E., Erhardt, J.B., Woody, E.A., Pratt, S.R., Getts, A., Kueser, T.J., Kolmer, J.W. and Sachs, M., 1986. Prevention of recurrent acute otitis media : chemoprophylaxis versus tympanostomy tubes. The Laryngoscope, 96(12), pp.1330-1334.

2. Kay, D.J., Nelson, M. and Rosenfeld, R . M . , 2 0 0 1 . M e t a - a n a l y s i s o f t y m p a n o s t o m y t u b e s e q u e l a e . Otolaryngology--Head and Neck Surgery, 124(4), pp.374-380.

3. M y e r I I I , C . M . , 2 0 0 1 . P o s t -tympanostomy tube otorrhea. Ear, Nose & Throat Journal, 80(6), p.4.

4. Post, J.C., 2001. Candidate's thesis: direct evidence of bacterial biofilms in otitis media. The Laryngoscope, 111(12), pp.2083-2094.

5. Prime, K.L. and Whitesides, G.M., 1991. Self-assembled organic monolayers: model systems for studying adsorption of proteins at surfaces. Science (New York, NY), 252(5009), pp.1164-1167.

6. Björling, M., Linse, P. and Karlström, G., 1990. Distribution of segments for terminally attached poly (ethylene oxide) chains. Journal of Physical Chemistry, 94(1), pp.471-481.

7. Jeon, S.I. and Andrade, J.D., 1991. Protein—surface interactions in the presence of polyethylene oxide: II. Effect of protein size. Journal of Colloid and Interface Science, 142(1), pp.159-166.

8. S p l e t e , H . ( 2 0 1 6 ) . W h e n G o o d Tympanostomy Tubes Go Bad : Pediatric News. [online] Pediatricnews.com. A v a i l a b l e a t : h t t p : / /www.pediatricnews.com/news/top-news/single-article/when-good-tympanostomy-

tubes-go-bad/1736f5abb3.html [Accessed 25 Apr. 2016].

9. Densert, B. and Densert, O. (1990). Method of ventilating the middle ear by means of a ventilation tube which can be applied in the tympanic membrane. US4971076A.

10. Berry, J. (2000). In vitro resistance to bacterial biofilm formation on coated fluoroplastic tympanostomy tubes. Otolaryngology - Head and Neck Surgery, 123(3), pp.246-251.

11. Ruohola, A., Meurman, O., Nikkari, S., Skottman, T., Salmi, A., Waris, M., Osterback, R., Eerola, E., Allander, T., Nies ters , H. , Heikkinen, T. and Ruuskanen, O. (2006). Microbiology of Acute Otitis Media in Children with Tympanostomy Tubes: Prevalences of Bacteria and Viruses. Clinical Infectious Diseases, 43(11), pp.1417-1422.

12. Stadelmann, W.K., Digenis, A.G. and Tobin, G.R., 1998. Physiology and healing dynamics of chronic cutaneous wounds. The American Journal of Surgery, 176(2), pp.26S-38S.

13. Brétagnol, F., Rauscher, H., Hasiwa, M., Kylián, O., Ceccone, G., Hazell, L., Paul, A.J., Lefranc, O. and Rossi, F., 2008. The effect of sterilization processes on the bioadhesive properties and surface chemistry of a plasma-polymerized p o l y e t h y l e n e g l y c o l f i l m : X P S c h a r a c t e r i z a t i o n a n d L 9 2 9 c e l l proliferation tests. Acta biomaterialia, 4(6), pp.1745-1751.

14. Med t ron ic Xomed Inc . , ( 2011) . ACTIVENT® ANTIMICROBIAL VENTILATION (TYMPANOSTOMY) TUBES. [online] Jacksonville, FL: Medt ron ic . Ava i l ab le a t : h t tp : / /www.bewusstlosigkeit.de/wcm/groups/mdtcom_sg/@emanuals/@era/@st/

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d o c u m e n t s / d o c u m e n t s /contrib_187560.pdf [Accessed 24 Apr. 2016].

15. "874.3930 - CFR - Code of Federal Regulations Title 21." 2009. 25 Apr. 2016 <https://www.accessdata.fda.gov/scripts/cd rh / c fdocs / c f c f r / c f r s ea r ch . c fm?fr=874.3930>

16. Anon, 2016. 501(k) Clearances. U.S. Food And Drug Administration. Available f r o m : h t t p : / / w w w . f d a . g o v /m e d i c a l d e v i c e s /p r o d u c t s a n d m e d i c a l p r o c e d u r e s /d e v i c e a p p r o v a l s a n d c l e a r a n c e s /510kclearances/ [Accessed April 24, 2016].

17. Anon, 2015. Overview of Device Regulation. U.S. Food And Drug Administration. Available from: http://w w w. f d a . g o v / m e d i c a l d e v i c e s /deviceregulationandguidance/overview/ [Accessed April 24, 2016].

18. Anon, 2016. Device Registration and L i s t i n g . U . S . F o o d A n d D r u g Administration. Available from: http://w w w. f d a . g o v / m e d i c a l d e v i c e s /d e v i c e r e g u l a t i o n a n d g u i d a n c e /h o w t o m a r k e t y o u r d e v i c e /registrationandlisting/ [Accessed April 24, 2016].

19. O f f i c e o f D e v i c e E v a l u a t i o n , Tympanostomy Tubes Submission Guidance for a 510(k) Premarket Notification, U.S. Department of Health and Human Services.

20. Anon, 2016. Premarket Notification 5 1 0 ( k ) . U . S . F o o d A n d D r u g Administration. Available from: http://w w w. f d a . g o v / m e d i c a l d e v i c e s /d e v i c e r e g u l a t i o n a n d g u i d a n c e /h o w t o m a r k e t y o u r d e v i c e /p r e m a r k e t s u b m i s s i o n s /

premarketnotification510k/default.htm [Accessed April 24, 2016].

21. Med t ron ic Xomed Inc . , ( 2002) . Antimicrobial tympanostomy tube. US6361526B1.

22. Universitaet Rostock, (2011). Tympanic membrane implant i.e. tympanostomy tube, for ventilation of tympanic cavity of patient during e.g. middle ear effusions, has small tube provided with outer flange between end-sided flange and end of through-hole. DE102010028705A1.

23. S u m m i t m e d i c a l u s a . c o m . ( 2 0 1 6 ) . Ventilation Tubes - PE Tubes - Ear Tubes - M y r i n g o t o m y - O t o l o g y - Tympanostomy. [online] Available at: http://www.summitmedicalusa.com/products/ent/otology/vent-tubes.php [Accessed 25 Apr. 2016].

24. Suyama Dental Laboratory Inc., (1997). Tympanostomy tube and method for producing the same. US5645584A.

25. Uptodate.com. (2016). Overview of t y m p a n o s t o m y t u b e p l a c e m e n t , postoperative care, and complications in children. [online] Available at: http://www.uptodate.com/contents/overview-of-t y m p a n o s t o m y - t u b e - p l a c e m e n t -postoperative-care-and-complications-in-children [Accessed 25 Apr. 2016].

26. Georgetown University, (1998). Tubular medical device. US5775336A.