A Novel In Vitro Method to Assess the Microbial Barrier Function of Tissue Adhesives Using Bioluminescence Imaging Technique

Abstract

Background. Tissue glues can minimize treatment invasiveness, mitigate the risk of infection, and reduce surgery time; ergo, they have been developed and used in surgical procedures as wound closure devices beside sutures, staples, and metallic grafts. Regardless of their structure or function, tissue glues should show an acceptable microbial barrier function before being used in humans. This study proposes a novel in vitro method using Escherichia coli Lux and bioluminescence imaging technique to assess the microbial barrier function of tissue glues. Different volumes and concentrations of E. coli Lux were applied to precured or cured polyurethane-based tissue glue placed on agar plates. Plates were cultured for 1 h, 24 h, 48 h, and 72 h with bioluminescence signal measurement subsequently. Herein, protocol established a volume of 5 μL of a 1 : 100 dilution of E. coli Lux containing around 2 × 10 7 CFU/mL as optimal for testing polyurethane-based tissue glue. Measurement of OD600nm, determination of CFU/mL, and correlation with the bioluminescence measurement in p/s unit resulted in a good correlation between CFU/mL and p/s and demonstrated good reproducibility of our method. In addition, this in vitro method could show that the tested polyurethane-based tissue glue can provide a reasonable barrier against the microbial penetration and act as a bacterial barrier for up to 48 h with no penetration and up to 72 h with a low level of penetration through the material. Overall, we have established a novel, sensitive, and reproducible in vitro method using the bioluminescence imaging technique for testing the microbial barrier function of new tissue glues.

Novel In Vitro Study to Assess Microbial Barrier Properties of Polyurethane-Based Tissue Adhesives in Comparison to the Gold Standard Dermabond®

“…The conventional in vitro test for evaluating the microbial properties of tissue adhesive is based on the production of organic acids by actively growing bacteria that results in a changing colour in the agar plate as an indication of a breach on the adhesive layer [ 27 ]. The herein-described in vitro test method used the bioluminescence imaging technique to detect and measure a certain wavelength of light produced by metabolically active growing bacteria as described before [ 29 ]. By employing an appropriate control group, a true penetration could be distinguished from random contamination, and a rational comparison would be obtained.…”

“…Escherichia coli Lux (kindly provided by Dr. Timo Schwandt) was cultured in sterile standard I nutrient broth medium (NB) (Carl Roth GmbH + Co. KG, Karlsruhe, Germany) containing 100 μ g/ml ampicillin (ampicillin sodium salt, Carl Roth GmbH + Co. KG, Karlsruhe, Germany) and incubated at 37°C in an incubator shaker with 190 rpm (Stuart Reciprocating Shaker SSL2, Cole-Parmer GmbH, Wertheim, Germany) as described before [ 29 ].…”

“…In addition, the method does not allow for studying scar formation, skin aging, and the effect of wound movement. However, due to the absence of an inherent immune system, our in vitro assessment might be more sensitive compared to in vivo models meaning that higher numbers of bacteria would be required to generate an infection in an in vivo model [ 27 , 29 ]. Therefore, in vivo models that provide direct analysis of a stimulus in the living human should be also considered.…”

“…Having a honey-like viscosity, they can be used to hold approximated skin edges of wounds in less invasive surgical incisions [ 28 ]. The purpose of the work described herein was to evaluate three different polyurethane-based tissue adhesives with different physiochemical properties for their microbial barrier function compared to Dermabond® using an in vitro method established in our previous study [ 29 ].…”

Design of biocompatible polylactic acid–polyethylene glycol combination for manufacturing biodegradable staplers

“…1,2 In terms of wound closure methods, staples, 3 sutures, 4,5 bandages, adhesives strips, 6 hydrogels, 7 and nanofibers can all be considered, each having their advantages and disadvantages. 8,9 For example, suturing non-linear wounds require special expertise, and the outcome is largely determined by the surgeon's expertise. It is still possible to get an infection despite the sutures' sterility since the spaces allow bacteria to colonize.…”