Ultra-Fast Glyco-Coating of Non-Biological Surfaces
"> Figure 1
<p>Schematic diagrams of the two primary blood group A function-spacer-lipid (FSL) constructs used in this paper. The upper schematic Atri-Ad-DOPE shows an FSL with a trisaccharide generic blood group A antigen and a short 2 nm adipate spacer while the lower schematic Atetra-CMG-DOPE shows a type 2 chain specific A tetrasaccharide FSL with a longer 7 nm carboxymethylglycine spacer. Both constructs have the same dioleoylphosphatidylethanolamine (DOPE) lipid tail.</p> "> Figure 2
<p>Photographic images of 20 representative coupons (see also <a href="#ijms-17-00118-t001" class="html-table">Table 1</a>) each spotted with 1 µL of Atri-Ad-DOPE (12 o’clock spot) and Atetra-CMG-DOPE (6 o’clock spot). Schematic diagram shows experimental layout. FSL spotted coupons were allowed to air dry then visualised by EIA using monoclonal anti-A and precipitating chromogenic substrate (purple).</p> "> Figure 3
<p>An example of microplate prepared with blood group <b>A</b> and <b>B</b> FSL constructs printed on paper (MG paper—cellulose esters) and prepared as a microplate to determine specificity of monoclonal reagents. Reprint from [<a href="#B8-ijms-17-00118" class="html-bibr">8</a>]. Copyright 2014 with permission from John Wiley and Sons. Alphanumeric characters appear when developed in the EIA reaction wherever the monoclonal reagent has bound to the FSL. Area outside of the printed area is the “internal” negative control and/or blank. In this example antibodies in co-ordinates <b>A1</b>, <b>A4</b>, <b>B3</b> have anti-A specificity, those in <b>A2</b>, <b>B1</b> and <b>B4</b> have anti-B specificity while those in <b>A3</b> and <b>B2</b> anti-A + B specificity. This EIA technique either with printed alphanumeric characters or spots of FSL constructs on a variety of different surfaces and optionally prepared as microplates was the basis for most experiments.</p> "> Figure 4
<p>Examples of 21 different surfaces reacted with Atri-Ad-DOPE. FSL constructs on most surfaces were inkjet printed as words identifying each surface material. The microspheres and glass fibre were coated with FSL construct by immersion, while a paintbrush was used to apply the FSL construct to gold foil. The blood group A FSL constructs bound to the various surfaces were visualised by EIA using monoclonal anti-A and a precipitating chromogenic substrate.</p> "> Figure 5
<p>Capture of red blood cells onto printed glyco-FSLs. Six different ABO related FSL constructs based on adipate and CMG spacers were printed onto five different surfaces. The upper row of images show the presence of these FSL constructs on surfaces using EIA and monoclonal antibodies directed against both the A and B antigens. The middle row of images show blood type specific binding of these same surfaces when reacted with monoclonal IgM anti-A and used to capture blood group A red cells. Of particular note is the poor EIA and strong red cell binding reaction of the silver membrane, highlighting the inability of some surfaces to show strong reactivity by the EIA method. The lower row of images are SEM 2000× magnifications of the edge of the printed areas, showing delineation between the printed blood group A FSL + IgM anti-A capture of blood group A red cells area and unprinted areas. It can be seen by its ability to immobilise RBCs that the monoclonal anti-A used had higher cross reactive affinity for the Forssman disaccharide (2FSad) than for the pentasaccharide (5FSad). This is expected as the Forssman antigen is a cross-reactive target of the anti-A reagent used, and the pentasccahride with a more complete Forssman structure will have less off-target binding [<a href="#B9-ijms-17-00118" class="html-bibr">9</a>].</p> "> Figure 6
<p>Attachment of blood group A red cells onto 20 µm polycarbonate microspheres coated with Atri-Ad-DOPE and IgM anti-A. Inset (<b>a</b>) shows light microscopy image at 1000× magnification while insets (<b>b</b>) & (<b>c</b>) show higher magnifications under SEM (post glutaraldehyde fixation). Although the main image and inset (<b>a</b>) appears to show cells only on the perimeter of the microsphere, this is an artefact due to the microscopic plane of focus. Varying the plane of focus reveals red cells evenly distributed over the microspheres. No binding occurred on controls.</p> "> Figure 7
<p>Incorporation of Atri-Ad-DOPE into nanofibres during the electrospinning process. Cellulose acetate (CA) and polyvinyl butyral (PVB) nanofibres were electrospun with Atri-Ad-DOPE and Btri-Ad-DOPE constructs included in the pre-spinning liquid polymers. Spun nanofibres were twisted into cords and then immunostained. Staining with monoclonal anti-A by the EIA method was able to demonstrate incorporation of the blood group A FSL construct into both nanofibres during the electrospinning process.</p> "> Figure 8
<p>Effect of molar concentration on the surface binding characteristics of Atri-Ad-DOPE and Atetra-CMG-DOPE when applied as a 1 µL spots on MG paper.</p> "> Figure 9
<p>Effect of application buffer pH on surface binding characteristics of Atri-Ad-DOPE and Atetra-CMG-DOPE when applied as a 1 µL spots (50 µmol/L) on paper. Columns show the time of contact in seconds of the FSL with the surface before being washed with PBS. The last clearly positive reaction (defined as even reactivity over the spot area) in each column is indicated by a yellow circle.</p> "> Figure 10
<p>Effect of application buffer ionic concentration on the surface binding characteristics of Atri-Ad-DOPE and Atetra-CMG-DOPE when applied as a 1 µL spots (50 µmol/L) on paper. Columns show the time of contact in seconds of the FSL with the surface before being washed away with PBS. The last clearly positive reaction in each column is indicated by a yellow circle.</p> "> Figure 11
<p>Effect of FSL application buffer (PBS, surfactant (Tw20), 70% ethanol) on the binding characteristics of Atri-Ad-DOPE and Atetra-CMG-DOPE when applied as a 1 µL spots (50 µmol/L) on paper, stainless steel and polyester film. Columns show the time of contact in seconds of the FSL with the surface before being washed away with PBS. The last clearly positive reaction in each column is indicated by a yellow circle.</p> "> Figure 12
<p>Effect of contact with human serum on the binding characteristics of Atri-Ad-DOPE and Atetra-CMG-DOPE when applied as a 1 µL spots (50 µmol/L) on paper, and exposed to serum for up to 24 h. After exposure to serum the presence of blood group A FSL constructs was determined by EIA with monoclonal anti-A. Serum concentration values relate to 100% being undiluted serum and 20% is serum diluted 1:5.</p> "> Figure 13
<p>Schematic diagrams of two additional blood group A function-spacer-lipid (FSL) constructs used in this paper. The upper schematic Atetra-Ad-DOPE shows an FSL with a tetrasaccharide blood group type 2 A antigen with an adipate spacer while the lower schematic is of an A trisaccharide with a cholesterol lipid tail (Atri-Chol).</p> "> Figure 14
<p>Elution profiles of four blood group A FSL variants printed on paper and exposed to 50%, 70% and 96% ethanol and methanol for 1 h. The FSL variants Atri-Ad-DOPE (3A0ad), Atri-Ad-sterol (3A0as), Atetra-Ad-DOPE (4A2ad) and Atetra-CMG-DOPE (4A2cd) when exposed to water (H<sub>2</sub>O) for 1 h all stained strongly by EIA. Exposure to 50% alcohol was effective at removing most constructs, but increasing concentrations of alcohol were less effective. Differences observed can be partially attributable to the type of lipid tail, spacer and size of the saccharide moiety. These results are probably only valid for this type of surface (paper).</p> "> Figure 15
<p>Detergent/surfactant stability of blood group A FSL on paper and nylon nanofibres. FSL variants Atri-Ad-DOPE (printed as FSL-Atri in image) and Atetra-CMG-DOPE (printed as FSL-Atetra in image) were printed on both paper and nylon nanofibres, exposed to deionised water (H<sub>2</sub>O), 70% methanol (MeOH), 5% Tween 20 or 5% Triton X-100 for 1 h, washed then immunostained by EIA.</p> ">
Abstract
:1. Introduction
2. Results and Discussion
2.1. Surface Variations
2.1.1. Surface Variations—Coupons
Surface | Alphabetical Listing of Materials Modified by Blood Group A Fsl Constructs |
---|---|
Metals | Aluminum, Copper, Gold, Nickel, silver, Stainless Steel (304), Stainless Steel (316L), Stainless Steel (347), Titanium |
Plastics/Polymers/Rubbers/Fibres (Alphabetical Order) | Acrylonitrile butadiene styrene (ABS), Cellulose acetate (transparency film), Cellulose acetate (nanofibres), Chlorinated polyvinyl chloride (CPVC), Chlorosulfonated polyethylene (CSPE, hypalon), Cotton, Ethylene propylene diene monomer (EPDM) rubber, Mixed cellulose esters, Natural rubber, Nitrile butadiene (NBR) rubber, Nitrocellulose, Poly(methyl methacrylate) (PMMA: Plexiglass), Polyamide (Nylon), Polyamide PA66 nanofibres, Polycarbonate, Polyetheretherketone (PEEK: Arlon 1330), Polyethylene terephthalate (PET: Polyester, Dacron), Polyethylene terephthalate glycol (PETG), Polyethylene UMHW, Polypropylene, Polystyrene, Polytetrafluoroethylene (PTFE), Polyurethane (high temperature polymer), Polyvinyl butyral nanofibres (PVB), Polyvinyl chloride (PVC), Polyvinylidene fluoride (PVDF), Regenerated cellulose, Silicone rubber, Silk, Silica gel S60 (TLC plate), Silica gel C18 (TLC plate), Viton rubber, wood (various) |
Other | Borosilicate glass, Concrete, Ceramic tile-glazed, Hydroxyapatite, Ceramic porcelain, Paper-24 varieties of coated and uncoated papers |
2.1.2. Surface Variations—Printing and Direct Application
2.1.3. Cell Adhesion to Printed FSL Constructs
2.1.4. Microsphere Binding
2.1.5. Nanofibre Incorporation
2.2. Binding Characteristics and Performance
2.2.1. Molarity
2.2.2. pH
2.2.3. Ionic Concentration
2.2.4. Effect of Surface and FSL Delivery Solutions
Solvent | Atri-Ad-DOPE | Atri-CMG-DOPE | Atetra-CMG-DOPE | |||
---|---|---|---|---|---|---|
Methanol | Ethanol | Methanol | Ethanol | Methanol | Ethanol | |
96% | + 1 | + | + | P 2 | + | P |
80% | + | + | + | + | + | + |
70% | + | + | + | + | + | + |
2.3. Stability
2.3.1. Serum
2.3.2. Alcohol
2.3.3. Detergent/Surfactant
3. Experimental Section
3.1. Synthesis of FSL Constructs
3.2. Surfaces
3.2.1. Disc Coupons
3.2.2. Membranes and Planar Surfaces
3.2.3. Electrospinning Nanofibres Incorporating FSL Constructs
3.3. Methods for FSL Coating Surfaces
3.3.1. Direct Application
3.3.2. Inkjet Printing
3.4. Microplate Assembly
3.5. Visualisation of Blood Group A FSL Constructs
3.5.1. Enzyme Immunoassay (EIA)
3.5.2. Cell Binding via Antibody Bound to FSL
3.6. Scanning Electron Microscopy (SEM)
4. Conclusions
Acknowledgments
Author Contributions
Conflicts of Interest
References
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Williams, E.; Barr, K.; Korchagina, E.; Tuzikov, A.; Henry, S.; Bovin, N. Ultra-Fast Glyco-Coating of Non-Biological Surfaces. Int. J. Mol. Sci. 2016, 17, 118. https://doi.org/10.3390/ijms17010118
Williams E, Barr K, Korchagina E, Tuzikov A, Henry S, Bovin N. Ultra-Fast Glyco-Coating of Non-Biological Surfaces. International Journal of Molecular Sciences. 2016; 17(1):118. https://doi.org/10.3390/ijms17010118
Chicago/Turabian StyleWilliams, Eleanor, Katie Barr, Elena Korchagina, Alexander Tuzikov, Stephen Henry, and Nicolai Bovin. 2016. "Ultra-Fast Glyco-Coating of Non-Biological Surfaces" International Journal of Molecular Sciences 17, no. 1: 118. https://doi.org/10.3390/ijms17010118