Polymers imprinted with three REG1B peptides for electrochemical determination of Regenerating Protein 1B, a urinary biomarker for pancreatic ductal adenocarcinoma

Abstract

Three peptides (each containing 13-18 amino acids) were synthesized and used as templates for molecular imprinting and epitope recognition of the regenerating Protein 1B (REG1B), which is one of the urinary biomarkers for pancreatic ductal adenocarcinoma (PDAC). Poly(ethylene-co-vinyl alcohol)s were employed as the host for molecular imprinting of the peptides. Following their preparation, the molecularly imprinted polymers (MIP) were examined by cyclic voltammetry. The electrochemical responses of a screen-printed gold substrate coated with the MIP were measured at a working voltage of 300 mV (vs. Ag/AgCl); the entire protein and the peptides gave similar responses at concentrations of <1.0 pg⋅mL⁻¹, with detection limits as low as 0.1 pg⋅mL⁻¹. Urine samples from healthy and PDAC patients were then analyzed by using this modified gold electrode, and the results are in agreement with data obtained with ELISA.

Epitope recognition of peptide-imprinted polymers for Regenerating protein 1 (REG1)

“…The effect of the concentration of peptides and REG1B on MIP-coated electrodes was then examined by cyclic voltammetry. As reported previously, [22] at lower concentration (less than 1.0 pg/mL), the response of REG1B to peptide MIP sensors is very close to that of peptides, suggesting that the entire protein can participate in electron transfer processes. At higher concentrations, the response falls off slightly for the protein compared to the imprinted peptide.…”

“…Peptides 2, 4, and 6 (from Reg1Α) have been used as templates in a previous study. [22] The current work studied peptides 1, 3, 5, and 7 (from Reg1Β), with a focus on peptide 7. The solubility of peptide 7 should be very good owing to its 6 charged (EDREDD) and 3 polar (NTY) amino acids.…”

“…Extraction of REG1B from E coli culture medium was performed using a KingFisher™ mL purification system (Thermo) with steps as follows: (1) MPIP particle collection: peptide http://tw.sinobiological.com/) was employed to examine the REG1B concentration; [22] the total protein was measured using a Protein Quantification Assay (740967.50, MACHEREY-NAGEL GmbH & Co. KG; http://www.mn-net.com/tabid/10972/default.aspx). All measurements in this work were carried out with at least two replicates; data are expressed as means and standard deviations.…”

Immunotherapy of Hepatocellular Carcinoma with Magnetic PD-1 Peptide-Imprinted Polymer Nanocomposite and Natural Killer Cells

“…Conventionally, the template molecule that is used in the imprinting of proteins is the whole protein, thereby providing high selectivity and sensitivity. Potential disadvantages of whole protein templates may include solubility limitations and price [19]. Thus, inexpensive proteins such as albumin [20], lysozyme [21], or hemoglobin were employed in model systems.…”

Immunotherapy of Hepatocellular Carcinoma with Magnetic PD-1 Peptide-Imprinted Polymer Nanocomposite and Natural Killer Cells

“…17 Conventionally, the template molecule that is used in the imprinting of proteins is the whole protein, to provide high selectivity and sensitivity. Potential disadvantages of whole protein templates include solubility limitations and price 18 for the preparation of MIPs. Therefore, some proteins such as albumin, 19 lysozyme 20 or hemoglobin have been employed as the model systems.…”