Reader and Industry Forum

Tearing the Tears Apart

By Yousra Djalal; Etty Bitton, OD, MSC, FAAO; & Lyndon Jones, Fcoptom, PHD, FAAO

The tear film has numerous roles including protecting and lubricating the ocular surface as well as serving as the eye's main refractive surface (Ohashi et al, 2006; Montes-Mico et al, 2010; Montes-Mico, 2007). Studying the physical and biochemical characteristics of the tear film in detail presents many challenges. This is primarily due to its very complex structure, which consists broadly of three “layers” that create various gradients of mucin, aqueous, and lipid throughout its depth; its thickness of only 3µm (King-Smith et al, 2000; Wang et al, 2006; Azartash et al, 2011); its dynamic nature, in which there is a constant change caused by blinks, evaporation, and drainage (Palakuru et al, 2007); and finally due to its very small volume of approximately 7µl (Mishima et al, 1966). Manufacturers of technologies involved in the study of the tear film have been faced with these challenges, and to date there is no single instrument that can address all of the complexities of this important biological fluid.

Figure 1. Glass capillary used to extract a tear sample from the lower tear meniscus.

Laser-Induced Breakdown Spectroscopy (LIBS)

One relatively new technology that may play a role in assisting with our understanding of the tear film is Laser-Induced Breakdown Spectroscopy (LIBS), which was first reported in the mid 1990s (Knopp et al, 1996; Vadillo et al, 1996). This technology vaporizes the sample of interest (liquid, solid, or gas) into its atomic elements (Vadillo and Laserna, 2004). The vaporized sample, termed a plasma, is composed of the atomic elements inherent to the sample (i.e. Ca⁺², Na⁺², Cl–). Each element in the plasma has a unique spectral emission and is subsequently detected and analysed by a spectrophotometer. The spectral line shown by LIBS depends on the intrinsic property of the element, its excitability, and its concentration (Thakur, 2007; Cremers and Radziemski, 2006), and this technique has been extensively used in analyzing the excipients of various samples such as tablets (Mowery et al, 2002; Fitzgerald et al 2005; Green et al, 2005).

The advantages of the LIBS technology are that there is no need for sample preparation (Bette and Noll, 2004) and, due to the precision of the laser, a sample as little as 13µl/cm² is sufficient (Sarkar et al, 2008), making it ideal for studying the ocular tear film.

Using LIBS, the tear film composition can be studied in its natural transparent state by collecting a small sample and running it through the LIBS instrument. LIBS also evaluates the tear film in its entirety, not each layer separately. More importantly, because only a small sample is needed, LIBS is particularly suited for studying dry eye patients, for whom tear sampling is more likely to be a challenge due to the reduced volume of tears present on the ocular surface.

Figure 2. The identification and quantification of sodium (Na) in a dried tear sample (green area) using LIBS.

LIBS and Contact Lenses

A stable and adequate tear film is essential to support comfortable contact lens wear, and several studies have reported that lens wear affects the tear film (Glasson et al, 2003; Faber et al, 1991; Maruyama et al, 2004; Chen et al, 2010). LIBS has been used in preliminary studies of contact lens materials (Jones, 2011) to analyze the elements of the lens and is a promising emerging technology in this field.

LIBS Challenges

The LIBS technology for analyzing the human tear film has its own set of challenges. For example, the recipient or container that holds the tears needs to be void of elements found in the tears. As such, a glass container or slide will be ill suited for the analysis, as the glass itself contains traces of calcium and magnesium, elements that are also found in the tears (Garg et al, 2006).

LIBS is a destructive, breakdown technology in which the tear samples are literally “torn apart” and consequently cannot be used for any other purpose.

LIBS is presently limited to a diagnostic laboratory setting, making the transportation of tear film samples to an outside source challenging, depending on the distance to the lab. Tear chemistry can be temperature-sensitive, so sample transportation can be an issue if the laboratory is too far away. Furthermore, technical expertise in spectrophotometry is required to run the LIBS and analyze the data. A close collaboration is needed between the lab and the investigators to ensure the integrity of the samples.

A Technique to Consider

In spite of the difficulties, LIBS is a promising technology in the study of tears, whether it be in relation to contact lens wear or in the realm of dry eye. CLS

For references, please visit www.clspectrum.com/references.asp and click on document #197.