Spectrometers

Trizol (Ambion Inc., Austin, TX, USA) reagent solution was added to the skin tissue, and total RNA was isolated. The amount and purity of total RNA were determined using 260 nm and 280 nm in spectrometers (Perkin Elmer). According to the manufacturer’s manual, total RNA (1 µg) was synthesized into cDNA using the Superscript™ III Reverse Transcriptase Kit (Bio-Rad, Richmond, CA, USA). The synthesized cDNA was mixed with SYBR Green supermix (Bio-Rad, Richmond, CA, USA). Its amplification was performed using the CFX Connect™ Real-Time PCR Detection System (Bio-Rad Laboratories, Inc., Hercules, CA, USA). Relative mRNA expression of TNF-α, interleukin (IL)-4, and IL-13 in the skin was assessed using the method of Ct comparison, and the expression level of the gene was normalized to that of the housekeeping gene β-actin. Table S1 lists the primers for the genes of interest.

The chemical functional group of ACN-g-ENR samples was assessed by Perkin-Elmer Frontiers Fourier-transform–infrared (FT-IR)/far-infrared (FIR) spectrometers, incorporated with the attenuated total reflection (ATR) accessory (Waltham, MA, USA), to indicate the grafting of ACN onto the ENR-25 structure. The analysis was done under a wavenumber from 650 to 4000 cm⁻¹ with a scan resolution of 2 cm⁻¹ at room temperature.