A-NARD – Trioscience

What is A-NARD?

A-NARD (Amplified Nucleic Acid Rapid Detection) is a relatively new diagnostic method that provides fast and reliable results through detection of the genomic material amplified with LAMP (Loop-associated Isothermal Amplification) method. It is based on rapid detection of the amplified genetic material with specific antibodies on lateral flow strips, with high accuracy. So, basically, the A-NARD method is the combination of two methods: LAMP and Lateral Flow tests. A-NARD method requires neither DNA isolation nor any additional equipment for the visualization of the results and gives comparable results to PCR with shorter time and lower costs, which are both important advantages for the laboratories. In addition, following the rapid and sensitive results obtained with the LAMP method, visualization of the results on lateral flow strip bands makes the method more reliable for the interpretation of results, which may indicate the presence of a life-threatening infection in the patient.

Advantages of A-NARD

A-NARD basically improves the visualization of the initial results obtained with LAMP method with the combination of a lateral flow assay; this yields higher sensitivity and specificity. Addition of NARD Buffer to the products obtained by LAMP can give highly accurate results on the lateral flow strip within minutes, either as positive or negative. A-NARD is relatively cheaper, faster and easier-to-apply compared to other methods used in the molecular diagnosis of infectious diseases.

What is LAMP?

LAMP (Loop-Mediated Isothermal Amplification) is a relatively new gene amplification method that provides a rapid, accurate, and cost-effective diagnosis of infectious diseases. It consists of three steps: The initial step, the cyclic amplification step, and the elongation step. PCR uses two primers, while the LAMP method works with at least six primers, which makes amplification possible over multiple points simultaneously. In addition, it is possible to identify multiple regions on DNA or RNA using LAMP, with higher sensitivity. During the execution of LAMP, a low number of nucleic acids are amplified in a short time; the change in the concentration of metal ions in the mixture and the rapid change in fluorescent stain creates a quick response associated with reaction accuracy.

Mechanism of LAMP

The LAMP primers are designed to define at least six regions around the target region (three primary forward and three primary reverses). The primer complexes, called FIP and BIP (inner), play a critical role in the loop formation and are connected to the nearest ends of the target region. The amplification from the inner primers results in loop formation at the ends of the DNA outline, which ensures the protection of the target gene. Two outer primers are designed for the distant ends of the target region. Outer primers are important for amplification to ensure the continuity of replication from the main sequence.

Advantages & Disadvantages

LAMP has essential advantages in nucleic acid amplification compared to other methods. Firstly, as it is an isothermal process, it does not need expensive ingredients such as PCR. Secondly, the loop structure formed in the target region and the adherence of the outer primers to the draft nucleic acid facilitate the production of the gene region. The target region is particular; it does not require any extra test procedures after amplification. In addition, any device that can provide a constant temperature (water bath, heated table, etc.) can be used in LAMP. RNA samples can also be studied easily with a reverse-transcriptase enzyme. LAMP is known to be more resistant than PCR in mixed samples that may contain enzyme inhibitors, such as blood and stool.
There are also disadvantages of LAMP method. Firstly, the primer design is the critical step in LAMP and should be done with high attention. Adverse folding of the primers may seldom occur, and cause dimer formation associated with false-positive results.

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