| Liquid crystal biosensor could detect analytes based on the orientation transition of liquid crystals as “sensitive element”,drived from specific bonding events of biomoleculars.This novel liquid crystal biosensing technology permited a label-free,highly sensitive and specific,low-cost,simple and fast detection for analytes and could be potentially applied in bioanalysis.In recent years,different analytical methods were developed for polypeptides detection,for instance,liquid chromatography-mass spectrometry,capillaryelectrophoresis and immunoassay.Most of these methods could sensitively and accurately detect polypeptides,however,it must also be mentioned that they need complicated pretreatment or derivatization,fluorescence labels for the samples and expensive laboratory equipment.In this paper,we carried out the following tasks through the combination of immunoassay and liquid crystal biosensor,considering the disadvantages of the ordinary methods for the detection of antimicrobial peptides.(1)The DMOAP and APTES with long and short alkyl chains were adopted to constructe the sensitive mixed self-assembled monolayers to decorate a plain glass slide substrate of liquid crystal biosensor,which can not only induce LCs vertical alignment to obtain a black background but also provide plenty of amino groups crosslinking with glutaraldehyde(GA)to immobilize biomolecules.The surface topography and wettability of liquid crystal biosening substrate were investigated by atomic force microscopy(AFM)and contact angle measurements,and the results indicated that DMOAP and APTES were successfully assembled on the surface of the glass slide substrate.The optical image of LC cell examzied by the polarizing microscope showed a uniform dark background,when the volume ratio of APTES/DMOAP was 3: 1 and the content of GA was 1%(v/v),which was benifit to biomolecules detection.This methodbased on the silanes self-assembled film is easy to operate,and can effectively orientate the liquid crystals orderly.(2)Based on the competitive immunoassay principle,we put forward a label-free liquid crystal immunosensor on the basics of orientation changes of liquid crystals for the detection of cecropin B using antigen cecropin B as a model.Cecropin B is a kind of small molecule antimicrobial peptides that has little effect on the orientation of the liquid crystal molecules,whereas,anti-cecropin B antibody with large molecular weight has significant influence on the alignment of the liquid crystal molecules.Therefore,in this work,we immobilized cecropin B on the surface of substrate modified with the mixed APTES/DMOAP self-assembled membrane,firstly,and then,observed the corresponding changes of the color and brightness of liquid crystal film,resulted from the competition binding with anti-cecropin B antibody between cecropin B antigen in the sample solution and cecropin B antigen on the sensing surface,which could sensitively and specifically detect cecropin B,as a result,the limit of cecropin B detection is as low as 0.1 ng/mL.(3)Based on the non-competitive immunoassay principle,a label-free liquid crystal immunosensor on the basics of orientation changes of LC molecules was reported for the detection of cecropin B.Anti-cecropin B antibody was immobilized on the surface of substrate modified with the mixed APTES/DMOAP self-assembled membrane,via coupling with glutaradehyde(GA)as a cross-linker.The specific binding event between cecropin B and anti-cecropin B antibody could induce homeotropic-to-tilted alignment transition of liquid crystal molecules.Ultimately,the specific detection of cecropin B could be achieved using the unique birefringent properties of liuid crystal moleculars,and as a result,the limit of cecropin B detection is as low as 50 ng/mL.Moreover,the average gray-scale intensties(GIs)of optical appearances were calculated to quantitatively analyse cecropin B concentrations.This study offers a simple,highly sensitive and specific,lable-free method for cecropin B detection. |
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