A novel, rapid, ultrasensitive diagnosis platform for detecting using restriction endonuclease-mediated real-time loop-mediated isothermal amplification.

INTRODUCTION

(, CA) is an essential invasive fungus in clinical diagnosis. Although several detection methods exist, none meet the need for early diagnosis. A rapid, sensitive, and specific diagnostic tool is crucial for effective prevention and control of infections.

METHODS

This study aimed to develop a new, rapid, and ultrasensitive diagnostic tool for detection based on restriction endonuclease-mediated real-time loop-mediated isothermal amplification (ERT-LAMP-CA). The ERT-LAMP-CA technology combines LAMP amplification, restriction endonuclease cleavage, and real-time fluorescence detection in a single reaction tube, which can complete a diagnosis of in a short time (approximately 1 h).

RESULTS

Herein, we developed the primer sequences required for ERT-LAMP-CA based on the ITS2 gene of and found that ERT-LAMP-CA limit of detection was approximately 500 ag/μL genomic DNA and can present negative results for non- templates. We tested sputum samples from 64 patients with suspected infections to validate ERT-LAMP-CA applicability in clinical sample testing and found that ERT-LAMP-CA was consistent with multiplex PCR-capillary electrophoresis.

DISCUSSION

In conclusion, ERT-LAMP-CA is a rapid, accurate, and sensitive assay with excellent potential for clinical and basic laboratory diagnosis and an efficient screening strategy.