INTRODUCTION:
In the last several years there has been an increasing demand for the development of infectious disease diagnostics which maintains test accuracy while reducing the time needed for detection at the point-of-care (POC) for the patient. Reducing the cost per assay as well as the equipment and end-user technical knowledge has been the driving force between the development of the lateral flow assay. This is primarily a qualitative colorimetric assay which can detect pathogen related proteins, nucleic acids, or antibodies. Here is a brief summary of several recent reviews for LFAs.
DISCUSSION:
Most conventional clinical diagnostics offers high specificity for the detection of specific pathogens. This specificity comes at the high cost of time, equipment and technical expertise to run these tests, not to mention the need to adhere to CLIA guidelines and testing regimens for a clinically approved assay. The LFA can overcome all of these obstacles. Basically, the LFA is a membrane (nitrocellulose or glass fiber substrate) which contains absorbed reagents which react to the applied sample in order to detect the target molecule (usually a pathogen related protein or specific antibody). However, other pathogen specific molecules can be applied to the LFA, such as aptamers (artificial nucleic acids which can selectively bind to target analytes), molecular beacons (a special DNA hairpin structure with an attached fluorophore which does not produce fluorescence in the absence of an analyte, but when a complementary DNA sequence (target nucleic acid) is present, the stem loop is opened and a fluorescent signal is observed), and finally, antibody bound colored magnetic particles. A review by Sajid et al.1Sajid, M., et al., 2015. Designs, formats and application of lateral flow assay: A literature review. J. of Saudi Chemical Society, 19, 689-705., discusses the details of the LFA construction. The use of aptamers may be advantageous since they are easy to generate/moderate, and can have a wide target range. In contrast, antibodies are not as stable and cannot be renatured, which has its limitations in developing new analytical methods2 Zhao, et al., 2018. State of the art: Lateral flow assay (LFA) biosensor for on-site rapid detection. Chinese Chemical Letters. 29, 1567-1577.. In addition to detecting clinically important pathogens, LFAs are used in a wide variety of assays for things such as, toxins, pesticides, metal ions, and drugs. Probably the most widely known over the counter LFA is the pregnancy test strip. Kozel et al.3Kozel, T.R., and Burnham-Marusich, A. R. 2017. Point-of-Care Testing for Infectious Diseases: Past, Present, and Future. J. Clin. Micro. 55, 2313-2320., discuss one of the most important advantages with the use of LFAs, namely, the waiver of CLIA guidelines. These authors list several POC tests using the LFA technology which waives the need for CLIA approval. The reason is that these tests are simple, qualitative and have a low risk of incorrect results. This includes serological LFAs for HIV-1/2 and hepatitis C virus. The detection of HIV antibodies from oral fluids using an LFA was given a CLIA-waived approval in 2012. In addition, viral antigen tests for Influenza A/B, respiratory syncytial virus (RSV), and Adenovirus have been CLIA-waived, as have several bacterial detection tests such as group A Streptococcus, H. pylori, Borrelia burgdorferi (Lyme disease), etc.
CONCLUSION:
The future of viral clinical assays using the lateral flow assay approach will be interesting to watch, but challenges exist. Most LFAs are simply qualitative, either positive or negative in their conclusion and monospecific (although multiplex assays continue to be explored/developed). This leads to their use in POC settings, and their ability to have CLIA waivers, but also limits their wider application in critical viral diagnostics until higher specificity and sensitivities can be achieved through improvement in the LFA technology.
