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P. E. King-Smith, J. J. Nichols, K. K. Nichols; Is Fluorescein Breakup Due to Quenching or Tangential Flow?. Invest. Ophthalmol. Vis. Sci. 2009;50(13):529.
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One assumption about fluorescein breakup is that it corresponds to "tangential flow" of the tear film away from the region of breakup, thus removing fluorescein from this region. We describe a method to test an alternative possibility that fluorescein is still present in the same amount (mass/area) and that evaporation causes "quenching" (reduced fluorescent efficiency) due to increase in fluorescein concentration.
Fluorescent efficiency is nearly independent of concentration at low concentrations, but falls rapidly within increasing concentration above a "critical concentration" of about 0.2% (Webber & Jones, 1986). Thus evaporation should reduce efficiency and fluorescence when the concentration of fluorescein in the tear film is high, but should have little effect for concentrations below 0.2%.
The figure compares fluorescence decay with high (1 uL of 10% instilled) and low (0.25 uL of 2%) fluorescein concentrations; the subject was a 68 year old white male with meibomian gland dysfunction who shows rapid, linear tear film thinning by interferometry. (Transient changes in the first 2 s after the blink are thought to be related to the upward movement of the lipid layer after a blink, causing tangential flow of the tear film.) Initial concentrations, just after the blink, were expected to be above and below the critical concentration respectively. Thus for the high concentration, increasing concentration due to evaporation should cause quenching and a rapid reduction in fluorescence. For the low concentration, there should be little quenching and fluorescence decay at first, until the concentration increases to above the critical concentration.
In this example, the observed fluorescence decays are consistent with the predictions based on increased concentration from evaporation causing quenching at relatively high concentrations. They are not consistent with tangential flow out of the measurement area which should cause similar decay curves for both high and low fluorescein concentrations. Thus a study of fluorescence decay using high and low fluorescein concentrations can be used to analyze the contributions of evaporation and tangential flow to tear film thinning and breakup.
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