Highly sensitive IRS based biosensor for the determination of cytochrome c as a cancer marker by using nanoporous anodic alumina modified with trypsin.

The determination of cytochrome c in the human serum sample is a regular medical investigation performed to assess cancer diseases. Herein, we used interferometric reflectance spectroscopy (IRS) based biosensor for the determination of cytochrome c. For this purpose first, the nanoporous anodic alumina (NAA) was fabricated. Then, the NAA pore walls were functionalized with 3-aminopropyl trimethoxy silane (NAA-NH2). Subsequently, the trypsin enzyme was immobilized on the NAA pore walls. The sensing principle of proposed IRS sensor to cytochrome c is based on a change in the intensity of the reflected light to a charge-coupled device (CCD) detector after digesting of cytochrome c by immobilized trypsin enzymes on NAA-NH2 into the heme-peptide fragment. The heme-peptide fragment then oxidized 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS) to green color ABTS·- anion radical in the presence of hydrogen peroxide. The generated green color ABTS·- anion radical solution adsorbed the white light and therefore the intensity of the reflected light from NAA to the CCD decreased. The decrease in the intensity of the white light had a logarithmic relationship with the concentration of the cytochrome c in the range of 1-100 nM. The limit of detections (LOD) for cytochrome c was 0.5 nM. The proposed biosensor exhibited high selectivity, sensitivity, and good stability.

Mesoporous silica nanotubes hybrid membranes for functional nanofiltration.

The development of nanofiltration systems would greatly assist in the production of well-defined particles and biomolecules with unique properties. We report a direct, simple synthesis of hexagonal silica nanotubes (NTs), which vertically aligned inside anodic alumina membranes (AAM) by means of a direct templating method of microemulsion phases with cationic surfactants. The direct approach was used as soft templates for predicting ordered assemblies of surfactant/silica composites through strong interactions within AAM pockets. Thus, densely packed NTs were successfully formed in the entirety of the AAM channels. These silica NTs were coated with layers of organic moieties to create a powerful technique for the ultrafine filtration. The resulting modified-silica NTs were chemically robust and showed affinity toward the transport of small molecular particles. The rigid silica NTs inside AAM channels had a pore diameter of

Aqueous two-phase extraction for protein recovery from corn extracts.

Corn has been used as an expression host for several recombinant proteins with potential for large-scale production. Cost-effective downstream initial recovery, separation and concentration remain a challenge. Aqueous two-phase (ATP) partitioning has been used to recover and concentrate proteins from fermentation broths and offers advantages for integration of those steps with biomass removal. To examine the applicability of ATP partitioning to recombinant protein purification from corn endosperm and germ, ATP system parameters including poly(ethylene glycol) (PEG) molecular weight (MW), phase-forming salt, tie line length (TLL), and pH were manipulated to control partitioning of extracted native proteins from each fraction. Moderate PEG MW, reduction of phase ratio, and added NaCl effected complete recovery of the hydrophobic model protein lysozyme in the top phase with ca. 5x enrichment and illustrates a favorable match of recombinant protein characteristics, expression host, and separation method. Furthermore, integration of protein extraction with the partitioning reduced the load of contaminating host proteins relative to the more traditional separate steps of extraction followed by partitioning. Performance of the integrated partitioning was hindered by endosperm solids loading, whereas for germ, which has ca. 35x higher aqueous soluble protein, the limit was protein solubility. For more hydrophilic model proteins (the model being cytochrome c), effective separation required further reduction of PEG MW to effect more partitioning of host proteins to the top phase and enrichment of the model protein in the lower phase. The combination of PEG MW of 1450 with 8.5 wt.% NaCl addition (Na(2)SO(4) as the phase-forming salt) provided for complete recovery of cytochrome c in the lower phase with enrichment of 9x (germ) and 5x (endosperm). As a result of lower-phase recovery, the advantage of simultaneous removal of solids is lost. The lower solubility of native endosperm proteins results in higher purity for the same enrichment.