HybriDetect LAMP Kits –
Molecular Biology Detection Systems from a Single Source

LAMP + Lateral Flow Assay (LFA)

This property is also the reason why so many scientists and developers combine the LAMP reaction with our lateral flow test strips (HybriDetect, HybriDetect 2T and HybriDetect Cassette). The combination is possible by using specially labeled primers for LAMP, which later interact with the test strip. For more detailed information on how this works, see the article Loop mediated isothermal Amplification (LAMP) & Lateral Flow. The test strips are used to evaluate the LAMP reaction, making them easy and straightforward to interpret even by untrained personnel. For these reasons, the test strips are the ideal add-on to LAMP as a point-of-need (PON) detection system.

Figure 1: Schematic representation of the LAMP-LFA test system

These kits are also available as hot start variants. The activity of the enzyme is blocked at ambient temperature due to molecular inhibition based hot start technology. The polymerase is only activated at 45–50 °C, which reduces non-specific amplification and the formation of primer dimers.

Below you will find an overview of the available HybriDetect LAMP kits (see table 1).

¹ The single-component kits contain the components relevant to LAMP (Bst polymerase, buffer with dNTPs, MgSO4, and Bst enhancer) as individual components for maximum flexibility.

²  The ready to use kits contain the components relevant to LAMP (Bst polymerase, buffer with dNTPs, MgSO4, and Bst enhancer) as ready to use reaction mix for easy and fast assay development.

Practical example: Detection of Alicyclobacillus spp.

We were able to demonstrate the compatibility of the LAMP kits with our HybriDetect test strips using the detection of Alicyclobacillus spp. as an example. Alicyclobacillus plays a major role in the spoilage of fruit juices and is therefore of economic interest to the food industry.

The HybriDetect LAMP Hot Start Kit was used to detect Alicyclobacillus. Primers for the LAMP reaction were designed using the free PrimerExplorer V5 program (see table 2). To make the amplificates visible on the strip, the FIB and BIP primers were labeled with biotin and FAM at their 5′ ends, respectively.

Table 2: Primers for the LAMP reaction using the free PrimerExplorer V5 program

The LAMP master mix consists of 12.5 µl of ready-to-use LAMP reaction mix, 9 µl of PCR water, 2.5 µl of primer mix (16 µM FIP and BIP as well as 2 µM F3 and F4 primers) and 1 µl of sample. The LAMP mixture was incubated for 20 minutes at 60 °C. The reaction was then stopped by heating to 80 °C for 10 minutes.

The reaction was evaluated using agarose gel electrophoresis (AGE) and lateral flow test strips, respectively. A 1.5% gel was used for the AGE. The evaluation with the HybriDetect – Universal Lateral Flow Test Strips was performed by applying 2 µl of the LAMP reaction and 80 µl of running buffer to the test strip. The evaluation was performed after 5 minutes.

If Alicyclobacillus spp. is present in a sample, part of the DNA is amplified, which can be evaluated by an AGE in the laboratory or an LFA at the PON. This becomes visible through corresponding bands in the gel or lines on the test strip (see figure 2).

To determine sensitivity, DNA was isolated from an Allicyclobacillus acidoterrestris culture and examined using the developed LAMP-LFA detection method. The assay was able to detect 50 pg of A. acidoterrestris DNA. The concentrations and parameters recommended by the kit were used. Further detection-specific adjustment of the LAMP parameters (reaction temperature, reaction time, primer concentration, etc.) can increase sensitivity even further (see figure 3).

It has also been shown that LAMP-LFA detection is capable of detecting various subspecies of Allicyclobacillus (A. acidoterrestris, A. Herbarius, A. acidiphilus, A. cycloheptanicus). Allicyclobacillus spp. can be specifically detected. Other organisms (P. polymyxa, L. brevis, Enterococcus sp., A. aceti, P. damnosus, M. cerevisiae, P. frisingensis), which are also frequently found in the food industry, are not detected (see figure 4).

Summary

Within a short period of time, LAMP-LFA detection was established using the LAMP kit and the HybriDetect Universal test strip. The total detection time from sample addition to result output is 30 minutes. Only a (mobile) heating block is required to perform the detection, which enables use at the point-of-need (PON).

This good compatibility is evident among our partners and customers. Between 2022 and 2024, our HybriDetect test strips were combined with a LAMP reaction in 20% of all applications.

Further Information

For more information regarding the combination of LAMP and HybriDetect check out the articles Loop mediated isothermal Amplification & Lateral Flow on our website and our summary about LAMP and lateral flow.

To learn more about LAMP primer design and assay optimization we highly recommend Lucigen’s webinar video.

Do you want to know how other scientists have combined LAMP and HybriDetect? Then have a look at our online literature database.