Optical clearing mechanisms characterization in muscle

Abstract

Optical immersion clearing is a technique that has been widely studied for more than two decades and that is used to originate a temporary transparency effect in biological tissues. If applied in cooperation with clinical methods it provides optimization of diagnosis and treatment procedures. This technique turns biological tissues more transparent through two main mechanisms — tissue dehydration and refractive index (RI) matching between tissue components. Such matching is obtained by partial replacement of interstitial water by a biocompatible agent that presents higher RI and it can be completely reversible by natural rehydration in vivo or by assisted rehydration in ex vivo tissues. Experimental data to characterize and discriminate between the two mechanisms and to find new ones are necessary. Using a simple method, based on collimated transmittance and thickness measurements made from muscle samples under treatment, we have estimated the diffusion properties of glucose, ethylene glycol (EG) and water that were used to perform such characterization and discrimination. Comparing these properties with data from literature that characterize their diffusion in water we have observed that muscle cell membrane permeability limits agent and water diffusion in the muscle. The same experimental data has allowed to calculate the optical clearing (OC) efficiency and make an interpretation of the internal changes that occurred in muscle during the treatments. The same methodology can now be used to perform similar studies with other agents and in other tissues in order to solve engineering problems at design of inexpensive and robust technologies for a considerable improvement of optical tomographic techniques with better contrast and in-depth imaging.

Software development for estimation of optical clearing agent’s diffusion coefficients in biological tissues

“…By estimating those parameters, it is possible to evaluate the necessary time to turn the tissue more transparent and to quantify that transparency as a function of the agent concentration used to treat the tissue [10]. The optical immersion clearing technique is a method initially described in 1997 [11] and presents great potential to be applied in clinical applications that use optical diagnosis or treatment methods [2].…”

“…Such treatment is completely reversible by natural rehydration in vivo and by assisted rehydration on ex vivo tissue samples [10]. The potential of the optical immersion clearing technique is very significant to be used with clinical procedures that apply light for diagnostic and treatment purposes [2,7,14].…”

“…At the same time, but with a slower pace, the agent in the solution begins to diffuse into the outer layers of the tissue [18]. The difference in viscosity properties and molecular size between water molecules and agent molecules provide that the diffusion of agent into deeper layers of tissue to create the RI matching mechanism takes a longer time [10].…”

“…This way, the individualization of the two mechanisms needs a refined method that produces accurate results. Such a method is described in literature [8,13] and we have recently proven its concept experimentally as described in our recent publications [7,10,18]. In those papers we describe how we have used thickness and collimated transmittance (Tc) measurements made from thin muscle samples under treatment with glucose and ethylene glycol (EG) solutions to estimate the diffusion time, diffusion coefficient and viscosity of these agents in muscle.…”

“…Our research has produced additional results. Those additional results were the estimated diffusion properties of water inside the muscle [10,18] and the amount of free water content in the natural muscle [7,10].…”