In recent years, with the rapid development of molecular biology techniques, nucleic acid-based diagnostic methods have been extensively established and widely applied in the laboratory testing of human diseases. Compared to other nucleic acid amplification techniques, isothermal amplification offers the advantages of being fast, efficient, and specific, and does not require specialized equipment. Therefore, since its emergence, it has been considered by many scholars as a detection method that could potentially rival PCR.

As a result, it is believed that the development and application of diagnostic instruments and kits based on isothermal amplification technology will represent a new direction for Point-of-Care Testing (POCT) in molecular diagnostics.

Introduction to the Principles of Mainstream Isothermal Amplification Technologies

1、Loop-Mediated Isothermal Amplification

(LAMP) Technology

Principle：Loop-mediated isothermal amplification (LAMP) utilizes Bst DNA polymerase, which has strand displacement activity. It designs four types of specific primers targeting six regions of the target gene, enabling efficient, rapid, and specific amplification of the target sequence under isothermal conditions. The amplification can be conducted at a constant temperature of 60-65°C, and within 15-60 minutes, a nucleic acid amplification of 10⁹ to 10¹⁰ times can be achieved.

Advantages and Applications：Due to its rapid reaction, high specificity, simple and easy-to-operate equipment, and results that are easy to interpret, isothermal amplification has been applied in the detection of pathogenic bacteria, parasites, viruses, diseases, and genetically modified products. It is also expected to be widely used in clinical infectious disease diagnosis, environmental monitoring, food safety, and other fields. It has become an ideal method suitable for molecular Point-of-Care Testing (POCT) platforms.

Classification：LAMP detection methods are classified into several types, including turbidimetry, pH indicator methods, fluorescent dye methods (such as NHB, calcein, SYBR Green, Syto, etc.), and fluorescent probe methods.

2、Recombinase Polymerase Amplification (RPA) Technology

Principle：The recombinase enzyme, when bound to primers, forms a protein-DNA complex that can search for homologous sequences within double-stranded DNA. Once the primers have located the homologous sequences, a strand exchange reaction occurs, leading to the formation and initiation of DNA synthesis, which exponentially amplifies the target region on the template. The displaced DNA strand binds to single-strand binding (SSB) protein to prevent further displacement. In this system, a synthesis event is initiated by two opposing primers, primarily relying on the activities of the recombinase, Bsu enzyme, and SSB protein. The technology can achieve rapid detection of the target of interest within 10-30 minutes at temperatures ranging from 37 to 42°C.

Advantages and Applications：RPA boasts high sensitivity, strong specificity, low dependency on specialized equipment, and the ability to integrate various detection formats. It is particularly suitable for point-of-care testing at the grassroots level and in field settings. RPA can be widely applied in in vitro diagnostics, animal diseases, food safety, biosafety, agriculture, and other fields.

Classification：RPA detection methods are categorized into several types, including gel electrophoresis, EXO probe method, Fpg probe method, lateral flow dipstick (LF-RPA), and flocculation analysis, among others.

Product Performance Showcase

01 pH Sensitive Dye RT-LAMP Reagents

pH Sensitive Dye RT-LAMP uses a visual dye as a sensitive indicator, under isothermal conditions of 60-65℃, to complete one-step RNA nucleic acid amplification using a constant temperature water bath or a standard PCR machine. In just 30 minutes, the change in color after the reaction can be used to determine whether a pathogen infection is present, with more intuitive results (positive is orange-yellow, negative is magenta). This method is suitable for rapid testing of large populations.

Recommended Products：

RT-LAMP pH Sensitive Dyestuff Kit（13906ES）
RT-LAMP pH Sensitive Dye Chromogenic Version Freeze-Dryable Kit（13920ES）

Performance Showcase：

1.RT-LAMP/RPA mix Selection Guide

Product Classification	RT-LAMP mix			RPA mix
Product Name	RT-LAMP Dye Assay Kit(UDG plus)	RT-LAMP pH Sensitive Dyestuff Kit	RT-LAMP pH Sensitive Dye Chromogenic Version Freeze-Dryable Kit	Quick Fuse Amplifier
Product Item Number	13762ES	13906ES	13920ES	16702ES
Product Category	Fluorescent Dye	pH Sensitive Dye	pH Sensitive Dye	Probe
Product Components	2 components Buffer、Enzyme mix	3 components Buffer、RT Enzyme、Bst Enzyme	4 components Buffer、RT Enzyme、Bst Enzyme、Lyoprotectant	4 components Buffer、Enzyme mix、Magnesium Acetate
Freeze-Drying Supported	No	Yes,without Lyoprotectant	Yes,Containing Lyoprotectant	No

 

2.LAMP/RT-LAMP Enzyme Selection Guide

Product Classification	Bst Enzyme		Reverse Transcriptase		UDG Enzyme
Product Name	Bst Plus	Lyophilized Bst Plus	Ⅲ Reverse Transcriptase	III Reverse Transcriptase, Glycerol-free	UDG	UDG,Glycerol-Free
Product Item Number	14402ES	14405ES	11111ES	11297ES	14455ES	14001ES
Enzyme activity	40 U/μL	60 U/μL	200 U/μL	600 U/μL	1 U/μL	1 U/μL
Product Components	3 components	2 components	2 components	single-component	single- component	single-component
Freeze-Drying Supported	No	Yes	No	Yes	No	Yes

 

3、RPA Enzyme Selection Guide

Product Classification	Bsu	T4 X	T4 Y	SSB	CK	EXO
Product Name	Bsu	T4 UvsX	T4 UvsY	gp 32	Creatine Kinase	Exonuclease III
Product Item Number	11078ES	11079ES	11080ES	11081ES	14502ES	14525ES
Enzyme activity	5 U/μL	2 μg/μL	2 μg/μL	5 μg/μL	2 μg/μL	100 U/μL
Product Components	single-component	single-component	single-component	single-component	single-component	2 components
Freeze-Drying Supported	No	No	No	No	No	No