Developing a fully validated lateral flow assay is a multi-step process that typically takes anywhere from 6 to 24 months depending on challenges and complexities of the assay. The various stages of the design cycle are outlined below. .
Phase I: Feasibility
Before allocating resources to developing an assay, the first step is to demonstrate that what you want to accomplish is feasible, both scientifically and financially. The process begins with developing initial product specifications. A product specification will include a description of what you are trying to detect (target analyte), where are you going to detect it (target matrix), and who you are detecting it in (intended use). These specifications narrow the scope of the project and make the path to demonstrating feasibility easier as there are now quantitative metrics. With these specifications in mind, a test can be designed to meet these specifications.
To begin feasibility studies requires both a test and samples. During the initial investigation, the availability of commercial reagents and samples will be determined. If antibodies are not available, a plan to develop a specific antibody may be required. Characterizing the cost and time associated with sourcing material is essential to writing a development plan. Some projects can end before they even get started due to limitations of commercially available material, or the cost to develop your own. Clinical samples are especially challenging. While there are many commercial vendors that offer sample collection under IRB, IEC, and/or MOH approved collection protocols, this is both time consuming and expensive. In the feasibility phase, typically at a minimum of 40 samples (based on the CLSI guideline EP9-A2: Method Comparison and Bias Estimation Using Patient Samples) will be required. Once procured, these samples can be used in parallel to a test method of your choosing to demonstrate that your intended use is feasible. If successful, the next step is development.
Phase II: Assay Development
The next step in the design process is development. An initial scoping stage will estimate the time required to complete development activities, determine quality requirements and provide estimates on time and cost to transfer to manufacture. A more detailed design specification document will be drafted. Assay optimization activities account for the majority of the time and effort in this stage with the time to complete being highly assay dependent. Cassettes, readers and sample preparation will be investigated during this stage. The end goal of development is a design freeze and the production of prototypes.
Phase III: Assay Verification
In the verification and validation stage, engineering lots are tested in replicate with clinical samples to determine sensitivity and selectivity of the assay. Limit of detection and limit of quantification values are determined statistically. Draft batch records for the manufacturing of the assay are written during verification.
Phase IV: Validation and Transfer to Manufacturing
In this final stage, batch records are finalized and released in preparation of transfer to manufacturing. Validation lots are produced for more comprehensive testing that will be utilized for regulatory activities such as submission of 510(k) applications to the FDA.
Now that you're familiar with the design cycle, the next step is to develop design specifications that will define the goals of the project and the metrics your assay needs to achieve.