Label-free single-substrate quantitative protein assay based on optical characteristics of cholesteric liquid crystals

Cholesteric liquid crystal (CLC) is characterized by the selective reflection in the Bragg condition for constructive interference. Instead of assembling a LC cell with a pair of glass substrates, only a single glass substrate modified with vertical alignment reagent and layered with a CLC film of 3.4 ± 0.2 μm in thickness was utilized in this study for the detection of bovine serum albumin (BSA). By fine-tuning the ratio of the thickness d to the helical pitch P of the CLC film, both qualitative and quantitative biosensing capabilities were demonstrated with an E7/R811 CLC (d/P close to but smaller than unity) and an E7/R5011 CLC (d/P > > 1) film, respectively. The E7/R811 film was in the unwound homeotropic state in the absence of BSA, but transitioned to the fingerprint state at 10⁻⁹ to 10⁻⁶-g/ml BSA and to the planar state between 10⁻⁵ and 10⁻²-g/ml BSA. Quantitative protein assay based on E7/R5011 CLCs was established through transmission spectrometric analysis, and detection sensitivity was optimized by adjusting the Bragg reflection wavelength. A limit of detection of 5.9 × 10⁻⁹ g/ml was achieved for the CLC film consisting of 2.0-wt% R5011 in the nematic host E7 and exhibiting a Bragg reflection wavelength of 700 nm. As a majority of clinical assays are performed on a single solid substrate, the CLC-based, single-substrate biosensing platform allows for better integration with current technologies. Besides, the protein array employed in this work can be further miniaturized and converted to a protein microarray format to facilitate high-throughput analysis.