Lateral flow assays

Lateral flow assays (LFAs), also known as immunochromatographic assays or rapid tests, are fast, easy-to-use and portable devices that can detect the presence of a target substance in a liquid sample without the need for specialised staff and expensive equipment. These tests are widely used in medical diagnostics for point of care testing (e.g. home testing) or laboratory use. Because of their compact size and reduced cost, LFAs are often referred to as Point-Of-Care Tests (POCT). The most common example of LFA is probably the home pregnancy test.

A typical lateral flow device is represented in the image below, and typically includes the following components:

  • Sample pad.
  • Conjugate pad, where the labelling reagent (e.g. antibody-conjugated EasyConjTM quantum dots) is dried. The labelling reagent should be released rapidly when the sample comes into contact.
  • Nitrocellulose membrane, where the antibodies are striped. The membrane is often laminated onto a plastic backing.
  • Wick/Absorbent Pad, controls the sample flow along the strip.
  • Plastic housing (cassette).
Schematic of a typical lateral flow assay device
Schematic of a typical lateral flow assay device

During a typical lateral flow test in a ‘sandwich’ format (see image below), the sample is added to the sample pad, the sample is added to the sample pad, then flows through to the conjugate pad, where releases the dried labelling reagent. The target and the labelling reagent start to mix and form a complex, migrating towards the antibodies immobilised on the nitrocellulose membrane. These immobilised antibodies capture the target-label complex, allowing for the formation of a line. If the target is present, a line will form where the test line is; if the target is not present (or below the limit of detection of the assay), a line will not form. The control line should always appear, as it signals that the sample and the reagents are flowing correctly. Any unreacted labels and excess reagent past the capture zone to the absorption pad.

Antibody-antigen reactions within a lateral flow assay
Antibody-antigen reactions within a lateral flow assay

The wicking rate is one of the most important characteristics of a nitrocellulose membrane, since it affects the reaction time between the antibodies and the target. Typically, the slowest the wicking rate, the more sensitive is the test.

Various types of labelling reagents can be used for the visualisation of the lines. The most commonly used material are gold nanoparticles (‘colloidal gold’) and latex beads. However, other labelling reagents have been developed in recent years, including enzyme conjugates, magnetic particles and fluorescent particles – such as quantum dots.

The lateral flow technology is particularly useful for the production of rapid and portable point-of-care tests. These tests have made a significant contribution to reducing the spread of infectious diseases, one recent example being COVID-19.

 

References

  • Merk Millipore. Rapid Lateral Flow Test Strips – Considerations for Product Development. 2013 (link)