For the last few decades, the immunochromatographic assay has been used for the rapid detection of biological markers in infectious diseases in humans and animals The assay, also known as lateral flow assay, is utilized for the detection of antigen or antibody in human infectious diseases. There are a series of steps involved in the development of these immuno-chromatographic test kits, from gold nano colloids preparation to nitrocellulose membrane coating (NCM). These tests are mostly used for qualitative assays by a visual interpretation of results. For the interpretation of the results, the color intensity of the test zone is therefore very significant. Herein, the study was performed on a malaria antigen test kit. Several studies have reported the use of gold nanoparticles (AuNPs) with varying diameters and its binding with various concentrations of protein in order to optimize tests. However, none of these studies have reported how to fix (improve) test zone band intensity (color), if different sized AuNPs were synthesized during a reaction and when conjugated equally with same amount of protein. Herein, different AuNPs with average diameter ranging from 10 nm to 50 nm were prepared and conjugated equally with protein concentration of 150 µg/mL with K_D = 1.0 × 10⁻³. Afterwards, the developed kits’ test zone band intensity for all different sizes AuNPs was fixed to the same band level (high) by utilization of an ultraviolet-visible spectrophotometer. The study found that the same optical density (OD) has the same test zone band intensity irrespective of AuNP size. This study also illustrates the use of absorption maxima (λ max) techniques to characterize AuNPs and to prevent wastage of protein while developing immunochromatographic test kits.

In the current virus infection and pandemic situation, the extremely convenient gold nanoparticles sensor (AuNPs) dependent antibody and antigen detection kits are very important assays for infection detection. Various laboratories and manufacturers across the globe are utilizing different kits development and manufacturing processes, but the development of kits and large scale manufacturing are two different things. A manufacturing process needs to be controlled for financial, quality and regulatory compliance. The best way to control the process is by statistical method. The process produces products that contain variables as critical quality attributes (dependent variables) are predicted over the processing time. There are distinct immune-chromatographic point to care diagnosis devices and kits available with different quality attributes like antigen-antibody proteins, aptamers, markers (nanoparticles, latex beads, and fluorescence) and devices performance attributes, etc. Such attributes add uncertainty to the durability of such instruments. The gold nanoparticles are one of the key attributes, which certainly defines the performance of kits (sensitivity, specificity and uniformity). To establishing the manufacturing process capability, the static tolerance intervals are critical for the synthesis of gold nanoparticles, protein conjugation, and nitrocellulose membrane coating. In the immune test kits, critical process parameters (independent variable) affect dependent responses can be controlled statistically by fitting purpose methodology. The manufacturing process is to be established in the state of engineering control by setting up the critical process parameters. Statistically, the manufacturers can predict and handle out-of-specification (OOS) results even if the process is inconsistent and unstable. This article provides scientific tools to the reader to determine whether or not a gold nanoparticles based immunechromatographic development process is statically capable of scale-up.

Immune‐chromatographic kits are being used since several years in the rapid detection of infectious diseases. It is also called the lateral flow technique, and is used for antigen or antibody detection. There are a series of steps involved in the development of these immune‐chromatographic test kits. Still, the preparation of gold nanoparticles (AuNPs) is an important quality variable for the immune‐chromatographic test kit sensitivity. The immune chromatographic test must be specific in detection for specific antigen and antibody; this implies that the test kit should not show a false result. Secondly, the test kit should be sensitive enough to give a readable result, and the intensity of the test line should increase or decrease with the concentration of an analytic sample. Various factors can influence the performance of a test. Temperature differences in AuNPs preparation can alter the assay kinetics and contribute to assay variability. Other factors such as assay components, manufacturing processes and reagent variation also contribute to assay precision and accuracy. It is important to note that assay reproducibility is the combined effect of individual sources of variability. The authors have synthesized AuNPs by immediately controlling the reaction temperature. Different batches of Malaria rapid test kit were developed and the test kit sensitivity was analysed. It was found that test kits designed with temperature‐controlled AuNPs sensor had reproducible uniformity in terms of batch to batch sensitivity than AuNPs synthesized by conventional Turkevich and Fern process.

Humanity has been continuously threatened by epidemics and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has shown greater epidemic potential. According to the World Health Organization, measures such as rapid diagnosis, immediate isolation, and precautionary contact tracing are key tools of epidemic control. The method of detection or testing is critical in this epidemiological control, where SARS-CoV-2-positive cases are increasingly growing, leading to community infection. Immunochromatographic test kits have been described for the diagnosis of various infectious diseases because of their rapid scalability, convenient use, and prompt validation. The benefits of the immunochromatographic test kits include evaluation of the sample in approximately 20 minutes, lower cost per sample, and simple directions to use. Such test kits are composed of an uncut sheet structure that is protein conjugated, labeled with markers, and reagents coated in a nitrocellulose membrane. The need for thorough vetting is a major concern for the Central Drugs Standard Control Organization, Indian Council of Medical Research and other regulators, as more businesses are rushing to produce serologic test kits for SARS-CoV-2 worldwide. Initial reports of 86%−89% sensitivity and 84.2%–98.6% specificity were reported for these kits. Nonetheless, many national reports show variability in test accuracy significantly among various commercial suppliers. The virus's more recent mutated strains (B.1.1.7, B.1.617, and B.1.351) have also raised concerns about their detectability using these test kits. In India, manufacturers are developing rapid test kits to detect SARS-CoV-2 by importing pre-antigen or antibody-coated uncut sheets from vendors and then cutting them into strips. Such sheets also have problems with specificity and are expensive. The possibility of developing kits with an indigenous protein coating and conjugation to detect antigens and antibodies needs to be explored by Indian researchers. This communication describes techniques to develop precise rapid test kits for detecting SARS-CoV-2.