Abstract
A new molecular imprinting technique was developed for molecularly imprinted polymer particles (MIPs). Particles were synthesized using organic silane chemistries by a sol-gel process, where the relative amount of active monomers was complementary matched to the relative amount of surface charges of the West Nile antibody template. Synthesized MIPs showed specific binding to affinity purified polyclonal West Nile antibodies (WNA) with a loading capacity of 80 µg/mg, while MIPs absorbed non-specific proteins at a loading capacity of 28 µg/mg. A dissociation constant of Kd=57.45 μM was measured from the binding isotherms. MIPs selectively absorbed 27 times more WNA than either albumin or immunoglobulin, while MIPs absorbed 16 times more WNA than non- imprinted particles (NIPs). Finally, fluorescently labeled MIPs were incubated in a high bind 96 well plate previously loaded with template, albumin, or immunoglobulin as an immunoassay test. Fluorescent MIPs significantly bound more to wells with WNA than any other control. Thus, the development of new affordable and robust immunoassays with MIPs would be possible in the future.
Highlights
There is a significant demand for robust and stable receptor molecules that can mimic biological molecules, such as antibodies [1]
Tetraethyl orthosilicate (TEOS), 3-aminopropyl triethoxysilane (APS), phosphate buffered saline (PBS), succinic anhydride, ammonium hydroxide, sodium hydroxide, and ATTO 495 NHS ester were purchased from Sigma-Aldrich, all reagent grade
Bovine serum albumin (BSA) and bovine gamma globulin (BGG) standard ampules were purchased from Thermo Scientific. 4-(2-Hydroxyethyl)piperazin-1-ylethanesulphonic acid (HEPES) was purchased from VWR, reagent grade
Summary
There is a significant demand for robust and stable receptor molecules that can mimic biological molecules, such as antibodies [1]. Relying only on natural recognition molecules has limited the uses and capabilities of many aspects of health sciences due to product expense and stability. These limitations have impacted low resource areas where product expense and limited cold-chain makes antibody-based diagnostics tough to implement. The absence of diagnostic tests limits disease treatment based their clinical symptoms and the local prevalence of the disease. While this method is generally effective, unnecessary or inadequate treatment may be administered. In low-resource settings, lateral flow assays are used because of their speed, simplicity, and relatively low cost. Molecular imprinting has been proposed to fabricate robust immunoassays [6,7,8]
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