Cas12a and Lateral Flow Strip-Based Test for Rapid and Ultrasensitive Detection of Spinal Muscular Atrophy.

Chunhua Zhang,Zhiqing Hu,Desheng Liang,Miaojin Zhou,Lingqian Wu,Miaomiao Chen,Zhuo Li

doi:10.3390/bios11050154

Abstract

Spinal muscular atrophy (SMA) is characterized by severe lethality and irreversible progression. Early diagnosis of SMA is of more practical significance with the emergence of effective therapy. However, existing techniques to identify SMA patients rely on cumbersome instruments, hindering their accessibility and application. An SMA-Cas12a-strip assay was developed with the integration of Cas12a-based nucleic acid detection, isothermal amplification, and lateral flow strip. The analytical performance of the assay was assessed with clinical samples. To explore its extensible utility, various specimens were tested. Validated with 168 clinical samples, the sensitivity and specificity of the SMA-Cas12a-strip assay were both 100%. The minimum detectable concentration of genomic DNA containing the target gene achieved 526 aM. The assay was compatible with specimens from several sources, and the turnaround time could be within 1.5 h. We developed a simple, cost-effective, and highly sensitive and specific assay to detect SMA patients. With little and field-portable equipment, the assay holds great promise in the detection of SMA patients, particularly in low-resource regions.

Highlights

Spinal muscular atrophy (SMA) is an autosomal recessive neurodegenerative disorder and is one of the most common genetic diseases leading to infant death, with a reported incidence of 1 in 6000–10,000 live births and a carrier frequency of 1 in 50 in the general population [1,2]
With the integration of Cas12a-based nucleic acid detection, isothermal amplification, and a lateral flow strip, we aimed to establish a method with convenience and low costs to detect the homozygous deletion of Survival motor neuron 1 gene (SMN1) exon 7 in SMA patients
Establishment of the SMA-Cas12a Assay with a Fluorescence Probe

Summary

Introduction

Spinal muscular atrophy (SMA) is an autosomal recessive neurodegenerative disorder and is one of the most common genetic diseases leading to infant death, with a reported incidence of 1 in 6000–10,000 live births and a carrier frequency of 1 in 50 in the general population [1,2]. SMA is featured by the irreversible degeneration of motor neurons in the spinal cord, which leads to progressive muscle weakness and respiratory failure. Survival motor neuron 1 gene (SMN1) is the causative gene of SMA [3], and more than 95% of SMA patients have a homozygous deletion of the SMN1 exon 7 [4,5]. The survival motor neuron 2 gene (SMN2) is highly homologous to SMN1 in humans, SMN2 cannot fully compensate for the loss of SMN1 [6]

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Conclusion