Impact of membrane pore structure on protein detection sensitivity of affinity-based immunoassay

Abstract

Understanding a membrane's morphology is important for controlling its fi nal performance during protein immobilization. Porous, symmetric membranes were prepared from a polyvinylidene fl uoride/N-methyl-2-pyrrolidinone solution by phase inversion process, to obtain membrane with various microsized pores. The concentration and surface area of aprotein dotted on the membrane surface were measured by staining with Ponceau S dye. The dotted protein was further scanned and analysed to perform quantitative measurements for relative comparison. The intensity of the red protein spot and its surface area varied depending on the membrane pore size, demonstrating the dependence of protein immobilization on this factor. The membrane with the smallest pore size (M3) showed the highest protein spot intensity and surface area when examined at different protein concentrations. An increase in the applied protein volume showed a linearity proportional trend to the total surface area, and an uneven round dot shape was observed at a large applied volume of protein solution.

Thermodynamic Studies of Bovine Serum Albumin (BSA) Adsorption on Nylon Membrane

“…The porosity (pore volume) value obtained from the wetting and weighting method is 74.2820±0.0411 %, thus, the nylon membrane sample was considered to have high membrane porosity. Theoretically, a combination of smaller pored membrane with high porosity would be desirable as a potential capture agent, as it would be expected to offer larger surface area for protein adsorption [12], [6]. Besides measuring the amount of protein adsorbed on membrane interface, it is important to evaluate the possible interaction involved between the protein molecule and membrane to ensure the robustness and flexibility of the developed diagnostic kits in real application.…”

“…The images were then analyzed using ImageJ for the pore size distribution. The measurement of porosity by using wetting and weighting technique has been discussed elsewhere [6], with specific gravity of water (1.0 g/cm3) and specific gravity of nylon (1.12 g/cm3). Then, for the analysis of functional groups, the nylon membranes were cut into 20 mm diameter and examined by using ATR-FTIR (Spectrum One, Perkin Elmer, US) at 4 cm-1 resolutions at 45˚ incident angle.…”

“…Usually, diagnostic kits are functionally based on the concept of immunoassays, such as those in the initial screens of typhoid, malaria and tuberculosis. Another example would be the very commonly available pregnancy test strip [6]. The assembly of the diagnostic-kit can be divided into two parts; the immunoagents extraction and membrane selection.…”

Suppression of Coffee-Ring Effect on Nitrocellulose Membrane: Effect of Polyethylene Glycol

“…The deposition of protein particles in Figure 5(a) is mostly concentrated at the edge of the spot, whereas in (b), the evaporation is evenly throughout the spot, resulting in a more uniform and dark appearance. It also shows that the protein particles do not accumulate as much at the edge of the spot as at the earlier spot (Ahmad et al, 2016a). However, the CRE produced was not significant enough in Figure 5(b), even though naked eye can be observed the ring-like effect.…”

Suppression of Coffee-ring Effect (CRE) in the Development of Low-cost Diagnostic Kit

“…Typically, the deposited protein or immunoagent on the membrane surface is in a lower concentration. If the working mechanism of the diagnostic kit is based on dot-blotting, a lower concentration of deposited protein or immunoagent often causes an unequal distribution of solute known as the coffee-ring effect (Ahmad et al, 2016b). The coffee-ring effect (CRE) is a ubiquitous phenomenon that commonly occurs in our daily lives.…”