Organophosphorous pesticide (OP) contamination could have serious adverse effects on human health and the environment. Due to the toxicity of OPs and the threat presented by their accidental or intentional release in populated areas, the determination and monitoring of these OPs in food products and environment is of great importance. OPs are present in very small quantities and therefore, methods for their detection need to be highly sensitive and selective. Here, we aimed to develop a simple and selective aptamer-based colorimetric assay for the detection of omethoate, which is one of the commonly used OPs. The principle of the assay is that single-stranded DNA (ssDNA)-wrapped gold nanoparticles (AuNPs) are resistant to salt-induced aggregation. By employing an “artificial antibody” organophosphorous pesticide-binding aptamer (OBA) as the recognition element, aptamer-wrapped AuNPs (Au-apta) show high selectivity towards omethoate, resulting in the disconnection of aptamers from AuNPs and the aggregation of AuNPs. As there is a significant color change from the interparticle plasmon coupling during the aggregation of AuNPs, the established assay showed good linearity between 0.1 and 10 μmol/L, with a low detection limit of 0.1 μmol/L. Other OPs such as profenofos, phorate, and isocarbophos would not interfere with the detection of omethoate despite having similar structures. Thus, the colorimetric method shows potential for use in the detection of omethoate in real soil samples.

•PA cross-linked recombinant human collagen–chitosan scaffold.•Fabrication in one-step lyophilization with neutralization.•HPLC detection of RHC loss rate•HUVEC proliferation and migration in scaffold•Angiogenesis associated gene expressions were increased in scaffold cell culturing.Porous scaffolds with appropriate pore structure, biocompatibility, mechanical property and processability play an important role in tissue engineering. In this paper, we fabricated a recombinant human collagen-peptide (RHC)–chitosan scaffold cross-linked by premixing 30% proanthocyanidin (PA) in one-step freeze-drying. To remove the residual acetic acid, optimized 0.2 M phosphate buffer of pH 6.24 with 30% ethanol (PBSE) was selected to neutralize the lyophilized scaffold followed by three times deionized water rinse. Ninhydrin assay was used to characterize the components loss during the fabrication process. To detect the exact RHC loss under optimized neutralization condition, high performance liquid chromatography (HPLC) equipped size exclusion chromatography column was used and the total RHC loss rate through PBSE rinse was 19.5 ± 5.08%. Fourier transform infrared spectroscopy (FT-IR) indicated hydrogen bonding among RHC, chitosan and PA, it also presented a probative but not strong hydrophobic interaction between phenyl rings of polyphenols and pyrrolidine rings of proline in RHC. Further, human umbilical vein endothelial cell (HUVEC) viability analyzed by a scanning electron microscope (SEM) and acridine orange/ethidium bromide (AO/EB) fluorescence staining exhibited that this scaffold could not only promote cell proliferation on scaffold surface but also permit cells migration into the scaffold. qRT-PCR exhibited that the optimized scaffold could stimulate angiogenesis associated genes VEGF and CD31 expression. These characterizations indicated that this scaffold can be considered as an ideal candidate for tissue engineering.

•Fabrication of recombinant human collagen-chitosan scaffolds by freezing drying•Influence of freeze drying protocols on lyophilized scaffolds•Pore size, microstructure, porosity, swelling and cell viability were compared.•The optimized porous scaffold is suitable for cell (HUVEC) seeding.Pore architecture of 3D scaffolds used in tissue engineering plays a critical role in the maintenance of cell survival, proliferation and further promotion of tissue regeneration. We investigated the pore size and structure, porosity, swelling as well as cell viability of a series of recombinant human collagen-peptide–chitosan (RHCC) scaffolds fabricated by lyophilization. In this paper, freezing regime containing a final temperature of freezing (Tf) and cooling rates was applied to obtain scaffolds with pore size ranging from 100 μm to 120 μm. Other protocols of RHC/chitosan suspension concentration and ratio modification were studied to produce more homogenous and appropriate structural scaffolds. The mean pore size decreased along with the decline of Tf at a slow cooling rate of 0.7 °C/min; a more rapid cooling rate under 5 °C/min resulted to a smaller pore size and more homogenous microstructure. High concentration could reduce pore size and lead to thick well of scaffold, while improved the ratio of RHC, lamellar and fiber structure coexisted with cellular pores. Human umbilical vein endothelial cells (HUVECs) were seeded on these manufactured scaffolds, the cell viability represented a negative correlation to the pore size. This study provides an alternative method to fabricate 3D RHC–chitosan scaffolds with appropriate pores for potential tissue engineering.