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Lateral flow assays, also known as immunochromatographic assays or strip tests, are immunoassays that are used to detect the presence, or absence, of a target analyte in a sample. Since they are suitable for point-of-care testing, provide a result extremely quickly, and offer simple, user-friendly operation they are rapidly growing in popularity.
Lateral flow assays have traditionally relied on the use of antibodies that have been conjugated to colored detection moieties such as gold nanoparticles or latex beads. Colorimetric readouts allow rapid visual assessment of the assay result, however, despite the extensive and well-documented use of colorimetric detection methods, many researchers are instead turning to fluorescent labels such as fluorescent dyes, fluorescent proteins or Europium particles, since these offer several advantages:
Expedeon has created a wide range of products to help you simplify the development of your lateral flow assay, including kits to produce fluorescently-labeled reagents.
Our Lightning-Link® technology enables the direct labeling of antibodies, proteins, peptides, or any other biomolecule with free amine groups. The product range includes kits for labeling biomolecules with a wide range of fluorophores, covering the spectrum from UV to far infra-red, and is therefore ideally suited to the production of reagents for use in lateral flow assays which require an immunofluorescent readout.
The Lightning-Link® kits are extremely quick and easy to use. The conjugation reaction is initiated simply by adding the biomolecule to a vial of the lyophilized mixture which contains the label of interest and incubating. With no separation steps, 100% of the material is recovered; and since the bond which is formed is covalent, the resulting conjugates are highly stable.
Although it is possible to visualize many fluorescently-labeled antibody conjugates by eye (provided the antibody conjugate is present at a suitable concentration), it is more common to use a specialized strip reader for fluorometric detection. This should be fitted with the relevant filters for detection of the fluorophore and, if a multiplexed lateral flow assay is performed, the properties of each fluorophore should be carefully considered.
To view the properties of each of our fluorophores, including maximal absorption and maximal emission wavelengths, and extinction coefficient values, download our fluorescence reference card.
Our Conjugate Check&Go! kit is a dipstick lateral flow assay for confirming the successful conjugation of an IgG antibody to a colored label. It is compatible with antibody conjugates produced using many of the fluorescent Lightning-Link® antibody labeling kits, as well as with conjugates generated using colloidal gold, InnovaCoat® GOLD, and Latex Conjugation Kits. The kit is compatible with IgG antibodies from multiple species, provided that they have an affinity for either Protein A or G, which are immobilized at the Test line.
We have used Conjugate Check&Go! to test antibody conjugates produced using a wide range of our Lightning-Link® antibody labeling kits.
For lateral flow assays that require an even higher degree of sensitivity, we offer our Europium Conjugation Kit. These allow direct labeling of antibodies, proteins, peptides, or any other biomolecule with free amine groups to 200nm Europium (Eu) chelate microspheres. Our Europium particles have a specially treated surface, allowing the generation of highly stable, covalent conjugates which are resistant to aggregation and require no extensive pH optimization
Labeling with Europium particles provides up to 15-fold higher sensitivity compared with other particle-based assays, in part due to Europium’s large Stokes shift. Europium has a maximal absorbance of 365nm and a maximal emission of 610nm, with a wide excitation spectrum and a narrow emission spectrum. Since the emitted fluorescence has a long lifetime (µseconds), Europium can be used for time-resolved fluorescence.
We have tested antibody conjugates produced using our Europium Conjugation Kit in both dipstick and strip test lateral flow assay formats and evaluated the assay readouts to produce a qualitative result with a UV transilluminator, and a quantitative result with a lateral flow strip reader.
During a dipstick assay, the conjugated antibody is added directly to a solution containing the target analyte, to which it binds. The lateral flow test strip is then dipped into the solution, and the analyte and the detection reagent flow along the nitrocellulose membrane, towards the wicking pad. The analyte becomes bound at a Test line, which typically consists of a second specific antibody that has been immobilized on the membrane. A Control line, usually composed of an anti-species antibody (against the host species of the antibody conjugate), is included further along the strip.
The main difference between a dipstick assay and a strip test assay is that a strip test includes a conjugate release pad, which is a portion of the lateral flow assay in which the conjugated antibody has been dried. The sample is added to the sample application pad, from which it flows into the conjugate release pad. The analyte and the detection reagent then move along the nitrocellulose membrane, towards the wicking pad. The analyte becomes bound at a Test line and, as in the strip test, a Control line is usually included further along the strip.
We offer a wide range of products which are suitable for fluorescent detection in lateral flow, including kits for labeling antibodies with fluorescent dyes, fluorescent proteins or Europium particles, as well as our Conjugate Check&Go! Kit to confirm successful antibody conjugation. If you have any questions, please contact us.