Novel critical point drying (CPD) based preparation and transmission electron microscopy (TEM) imaging of protein specific molecularly imprinted polymers (HydroMIPs)

We report the transmission electron microscopy (TEM) imaging of a hydrogel-based molecularly imprinted polymer (HydroMIP) specific to the template molecule bovine haemoglobin (BHb). A novel critical point drying based sample preparation technique was employed to prepare the molecularly imprinted polymer (MIP) samples in a manner that would facilitate the use of TEM to image the imprinted cavities, and provide an appropriate degree of both magnification and resolution to image polymer architecture in the <10 nm range. For the first time, polymer structure has been detailed that clearly displays molecularly imprinted cavities, ranging from 5-50 nm in size, that correlate (in terms of size) with the protein molecule employed as the imprinting template. The modified critical point drying sample preparation technique used may potentially play a key role in the imaging of all molecularly imprinted polymers, particularly those prepared in the aqueous phase

A fully automated linear polyacrylamide coating and regeneration method for capillary electrophoresis of proteins

Surface modification of the inner capillary wall in CE of proteins is frequently required to alter EOF and to prevent protein adsorption. Manual protocols for such coating techniques are cumbersome. In this paper, an automated covalent linear polyacrylamide coating and regeneration process is described to support long-term stability of fused-silica capillaries for protein analysis. The stability of the resulting capillary coatings was evaluated by a large number of separations using a three-protein test mixture in pH 6 and 3 buffer systems. The results were compared to that obtained with the use of bare fused-silica capillaries. If necessary, the fully automated capillary coating process was easily applied to regenerate the capillary to extend its useful life-time

Pharmaceutical And Biomedical Applications Of Affinity Chromatography: Recent Trends And Developments

Affinity chromatography is a separation technique that has become increasingly important in work with biological samples and pharmaceutical agents. This method is based on the use of a biologically-related agent as a stationary phase to selectively retain analytes or to study biological interactions. This review discusses the basic principles behind affinity chromatography and examines recent developments that have occurred in the use of this method for biomedical and pharmaceutical analysis. Techniques based on traditional affinity supports are discussed, but an emphasis is placed on methods in which affinity columns are used as part of HPLC systems or in combination with other analytical methods. General formats for affinity chromatography that are considered include step elution schemes, weak affinity chromatography, affinity extraction and affinity depletion. Specific separation techniques that are examined include lectin affinity chromatography, boronate affinity chromatography, immunoaffinity chromatography, and immobilized metal ion affinity chromatography. Approaches for the study of biological interactions by affinity chromatography are also presented, such as the measurement of equilibrium constants, rate constants, or competition and displacement effects. In addition, related developments in the use of immobilized enzyme reactors, molecularly imprinted polymers, dye ligands and aptamers are briefly considered

Microfluidics: reframing biological enquiry

The underlying physical properties of microfluidic tools have led to new biological insights through the development of microsystems that can manipulate, mimic and measure biology at a resolution that has not been possible with macroscale tools. Microsystems readily handle sub-microlitre volumes, precisely route predictable laminar fluid flows and match both perturbations and measurements to the length scales and timescales of biological systems. The advent of fabrication techniques that do not require highly specialized engineering facilities is fuelling the broad dissemination of microfluidic systems and their adaptation to specific biological questions. We describe how our understanding of molecular and cell biology is being and will continue to be advanced by precision microfluidic approaches and posit that microfluidic tools - in conjunction with advanced imaging, bioinformatics and molecular biology approaches - will transform biology into a precision science