Preparation of ready-to-use, storable and reconstituted type I collagen from rat tail tendon for tissue engineering applications

Abstract

Collagen is a widely investigated extracellular matrix material with extensive potentials in the field of tissue engineering. This protocol describes a method to prepare reconstituted collagen that can be ready-to-use, storable and suitable for further in vitro and in vivo investigations. Type I collagen was extracted from rat tail tendons and processed in acetic acid solution to obtain sterile soluble collagen. At first, crude collagen was dissolved in acetic acid, then frozen at -20 degrees C and lyophilized to obtain a sponge, which could be stored at -80 degrees C. Lyophilized collagen was then dispersed in acetic acid to obtain a sterile solution of collagen at targeted concentrations. The whole low-cost process from the extraction to the final sterile solution takes around 2-3 weeks. The collagen solution, once neutralized, has the potential to be used to produce gels or scaffolds, to deposit thin films on supports and to develop drug delivery systems.

Tailoring Mechanical Properties of Collagen-Based Scaffolds for Vascular Tissue Engineering: The Effects of pH, Temperature and Ionic Strength on Gelation

“…Collagen was obtained from RTT and solubilized at 4 mg/mL in 0.02 M acetic acid according to a previously developed protocol [10]. Collagen gels were prepared on ice at two different pHs, with seven different salt concentrations corresponding to different ionic strengths, and gelled at three different temperatures.…”

“…Table 2. Results from the turbidity measurements as a function of pH (7,10), ionic strength (c1, c2, c3, c4, c5, c6, c7) and temperature (T1 = 4 °C, T1* = 4 °C and 21 °C after 38 days, T2 = 21 °C, T3 = 37 °C): Final absorbance for all the types of gels and t 1/2 for gels prepared at pH 10. The kinetics of gels prepared at pH 7 as a function of collagen concentration, temperature and ionic strength has already been widely discussed by several authors [23,24,30,31].…”

“…Table 3. CM and CSE as a function of ionic strength (c1, c2, c3, c4, c5, c6, c7), pH (7,10) and temperature (T1 = 4 °C, T2 = 21 °C, T3 = 37 °C) and expressed as mean ± SD. Tensile tests were conducted only on the types of gel that were considered the most interesting in regard to the improvement of the mechanical properties shown in compression.…”

“…Type I collagen can be obtained from the collagen fibers of rat-tail tendons (RTT) by solubilization in acetic acid [10]. This process results in the decomposition of collagen fibers and fibrils into small aggregates of collagen monomers dispersed in the acid solution.…”

Preparation and Characterization Urea-Solubilized Sol-Gel Type I Collagen and Its Possible Use in Applications

“…For this reason, an understanding of the structure of the whole collagen protein is still required [Orgel, et al, 2001]. However, this task is always hampered by the low quality of the collagen derived from conventional procedures, including neutral salt and acetic acid-extraction [Becker, & Timpl, 1972;Miller, et al, 1982;Chandrakasan, et al, 1976;Rajan, et al, 2007] (A comparison of the three procedures is shown in Fig. 1).…”