Decellularization of placentas: establishing a protocol

Biological biomaterials for tissue engineering purposes can be produced through tissue and/or organ decellularization. The remaining extracellular matrix (ECM) must be acellular and preserve its proteins and physical features. Placentas are organs of great interest because they are discarded after birth and present large amounts of ECM. Protocols for decellularization are tissue-specific and have not been established for canine placentas yet. This study aimed at analyzing a favorable method for decellularization of maternal and fetal portions of canine placentas. Canine placentas were subjected to ten preliminary tests to analyze the efficacy of parameters such as the type of detergents, freezing temperatures and perfusion. Two protocols were chosen for further analyses using histology, scanning electron microscopy, immunofluorescence and DNA quantification. Sodium dodecyl sulfate (SDS) was the most effective detergent for cell removal. Freezing placentas before decellularization required longer periods of incubation in different detergents. Both perfusion and immersion methods were capable of removing cells. Placentas decellularized using Protocol I (1% SDS, 5 mM EDTA, 50 mM TRIS, and 0.5% antibiotic) preserved the ECM structure better, but Protocol I was less efficient to remove cells and DNA content from the ECM than Protocol II (1% SDS, 5 mM EDTA, 0.05% trypsin, and 0.5% antibiotic).

Decellularization of placentas: establishing a protocol.

Biological biomaterials for tissue engineering purposes can be produced through tissue and/or organ decellularization. The remaining extracellular matrix (ECM) must be acellular and preserve its proteins and physical features. Placentas are organs of great interest because they are discarded after birth and present large amounts of ECM. Protocols for decellularization are tissue-specific and have not been established for canine placentas yet. This study aimed at analyzing a favorable method for decellularization of maternal and fetal portions of canine placentas. Canine placentas were subjected to ten preliminary tests to analyze the efficacy of parameters such as the type of detergents, freezing temperatures and perfusion. Two protocols were chosen for further analyses using histology, scanning electron microscopy, immunofluorescence and DNA quantification. Sodium dodecyl sulfate (SDS) was the most effective detergent for cell removal. Freezing placentas before decellularization required longer periods of incubation in different detergents. Both perfusion and immersion methods were capable of removing cells. Placentas decellularized using Protocol I (1% SDS, 5 mM EDTA, 50 mM TRIS, and 0.5% antibiotic) preserved the ECM structure better, but Protocol I was less efficient to remove cells and DNA content from the ECM than Protocol II (1% SDS, 5 mM EDTA, 0.05% trypsin, and 0.5% antibiotic).

Photoacoustic spectroscopy as a tool for determination of food dyes: comparison with first derivative spectrophotometry.

Photoacoustic spectroscopy (PAS) was applied as a method to quantify dyed food samples, and was compared with First Derivative Spectrophotometry (FDS). The dyes Brilliant Blue (B), Sunset Yellow (S) and Tartrazine (T), which are common food additives, were employed for the comparisons. Polyester-type Polyurethane (PU) foam was used for extraction of the dyes from a solution containing the food matrix. For the spectrophotometric determinations, the adsorbed dyes were recovered by using dimethylformamide. The PAS measurements were carried out directly on the PU foam. The PAS method showed greater sensitivity, with detection limits of 0.028 mg L(-1) and 0.086 mg L(-1) for S and T, respectively, in the S+T mixture, and of 0.012 mg L(-1) and 0.068 mg L(-1) for B and T, respectively, in the B+T mixture. The values of relative error obtained for all the dyes were small: approximately 0.3-3.6% for the spectrophotometer, and approximately 0.1-2.9% for the PAS method. The PAS technique can be applied to the determination of the selected dyes in commercial food products, with some advantages: it reduces the number of analysis steps, it is a "green" method with less chemical waste, a minimal sample amount is needed, and it is non-destructive.