Homing-peptide combined with malachite green based ELISA-like assay for highly specific and sensitive fibrin detection

Abstract

Sensitive, accurate and reliable fibrin detection is of great significance for evaluating the treatment effect of orthodontic implants. However, current fibrin detection methods lack enough sensitivity. We demonstrate here a novel fluorescence-based enzyme linked immunosorbent assay-mimicking method for fibrin detection. The sensing platform utilizes the homing peptide CREKA as a recognition molecule, enabling highly-selective fibrin binding onto the plate. Upon sample deposition, the CREKA-functionalized probe cross-links with fibrin via peptide–target interaction. Subsequently, rolling circle transcription (RCT) is initiated to synthesize malachite green (MG)-binding RNA aptamers, facilitating label-free fibrin detection through MG-derived fluorescence. Leveraging the high amplification efficiency of RCT, this assay achieves a detection limit of 0.01 fmol L⁻¹ and a wide detection range (from 0.01 fmol L⁻¹ to 5 pmol L⁻¹). Compared with the former method, the proposed method possesses a greatly elevated specificity by integrating the dual recognition of fibrin by antibodies and CREKA with split MG RNA aptamer induced signal generation. Considering the merits of the low limit of detection, high accuracy, and improved stability, the proposed method could be a promising tool for disease diagnosis.