Three-dimensional thin film coating


NEXTERION® Slide H

SCHOTT launched NEXTERION® Slide H as a dedicated slide surface for printing protein microarrays, as it is ideally suited for the covalent immobilization of peptides and proteins such as antibodies, antibody fragments, enzymes, or receptors. For many protein microarray applications NEXTERION® Slide H has proven to be a very attractive alternative to the commonly used nitrocellulose coated slide, especially where low background, or slide transparency are important considerations. Since its introduction, the slide coating has also been successfully used with amino-modified oligonucleotides, and has become the slide of choice for printing amino-linked glycan microarrays.
Carbohydrate arrays are a rapidly growing area of microarray research and NEXTERION® Slide H is an excellent choice for use in the rapid screening of carbohydrateprotein interactions. The permeable, polymer coating has a large immobilization capacity, and helps to preserve the native three-dimensional structure of complex bio-molecules, thus maintaining conformation and functionality.
NEXTERION® Slide H produces excellent signal-to-background ratios and an exceptionally wide dynamic range compared to conventional “two-dimensional” coatings through a unique combination of low non-specific binding characteristics, and high probe loading capacity. Even very low intensity signals, such as those obtained from low-abundance analytes, or weakly expressed genes can be reliably detected and quantified on NEXTERION® Slide H. The robust coating matrix is fully compatible with commercial microarray printers and scanners. Simple and robust protocols are available making NEXTERION® Slide H easy-to-use.


Overview

Type of coating Immobilization method Typical probes Ordering information
NEXTERION® product Barcode option Item number Slides per pack
Thin film 3-D polymer surface Amine reactive chemistry
Covalent binding
• Proteins
• Amino-linked glycans
• Amino-linked oligonucleotides
Slide H Laser 1070936 25
Key product features
• Ideal substrate for printing protein, amino-modified glycans or oligonucleotide microarrays
• High probe loading capacity
• Exceptionally wide dynamic range
• Very low non-specific binding characteristics
• Optimal preservation of native structure and biological activity of protein probes
• Compatible with all common microarray printers and scanners
Typical applications
• Proteome expression profiling
• Functional protein arrays
• Transcriptional profiling
• Biomarker discovery
• Detection of protein modifications
• Carbohydrate–protein interactions
• Bacteria serotyping
• Immunoassays
Suitable probe types
• Antibodies and antibody fragments
• Functional proteins such as enzymes or receptors
• Small protein fragments such as peptides
• Amino-modified oligonucleotides 16 to 70 mers
• Amino-linked glycans
• Cells

Immobilization chemistry

The coating on the SCHOTT NEXTERION® Slide H consists of a cross-linked, multi-component polymer layer activated with N-Hydroxysuccinimide (NHS) esters to provide covalent immobilization of amine groups. All NEXTERION® microarray slides are manufactured from a high quality, low-fluorescence glass coated with low-fluorescence coatings. However, the non-specific binding of assay components still remains an important contributor to the off feature background for many microarray applications. For most types of slide coatings the post-print processing protocol involves a method of adsorptive blocking to reduce non-specific binding, however, these procedures are difficult to perform in a consistent manner.
Immobilization chemistry NEXTERION hydrogel coated microarray slides
The NEXTERION® Slide H coating has been engineered to exhibit a very low intrinsic non-specific background without the need for blocking. This is achieved by using a polymer based layer that is extremely resistant to nonspecific binding. The polymer coating has a three-part structure; NHS-ester reactive groups are attached to the cross-linked hydrophilic polymer layer via long, flexible spacers. The terminal amino group of amino-modified nucleic acids and glycans react immediately and irreversibly with the NHS-ester groups to form a covalent bond. Proteins and other bio-molecules bind via surfaceexposed amine-groups. The flexible spacers tether the immobilized bio-molecules in a quasi-liquid environment that maintains the protein specificity and chemical conformation. The high accessibility of the tethered bio-molecules facilitates interactions with their binding targets in solution.


Product details

Highly reproducible coating
NEXTERION® Slide H is fabricated using a proprietary thin-film deposition process optimized by SCHOTT to produce a uniform and reproducible polymer coating on one side of a high quality borosilicate glass slide. All slides are individually examined for physical defects and the presence of particles before and after coating. The surface is applied in a tightly controlled, class 100 clean room facility, resulting in coated slides with highly uniform surface properties and low auto-fluorescence.

Excellent spot morphology and signal-to-background ratios
NEXTERION® Slide H provides excellent spot morphologies and reproducible spot sizes over a wide range of probe concentrations for protein, oligonucleotide and other bio-molecule microarray applications.
Spot morphology NEXTERION hydrogel coated microarray slides




Figure shows scanned images of NEXTERION® Slide H when evaluated against the market leading threedimensional competitor slide in an anti-IgG/IgG interaction study using a range of probe concentrations.
Signal-to-Background ration NEXTERION Slide H
NEXTERION® Slide H
Signal-to-background-ratio of NEXTERION hydrogel coated slides
Leading 3D slide
Epoxysilane slide
Figure illustrates the superior signal-to-background ratios on NEXTERION® Slide H when compared to the market leading three-dimensional slide, and a conventional two-dimensional coating. This was demonstrated in two-color hybridization experiments following the recommended protocols, using amino-modified oligonucleotide probes.

Packaging and storage
NEXTERION® Slide H are packaged in chemically stable plastic boxes and sealed under an inert atmosphere. The slides are ready-to-use from the box, and are stable for 12 months in the sealed packaging when stored at – 20 ºC.


Format
NEXTERION® Slide H is available in packs of 25 slides with code 128 barcodes enabling automated sample tracking. The identical coating is also available in a 16-well and 48-well slide format. For further information refer to the section “Multi-well formats”.

Compatible reagents

Protocol step Recommended NEXTERION® products Alternatives Recommended concentrations
Spotting (Protein) NEXTERION® Spot PB 150 mM phosphate, pH 8.5,
0.01% sarcosyl or Tween20©
Spotting (DNA) NEXTERION® Spot 300 mM sodium phosphate (pH 8.5) containing 0.005 % Tween20® and 0.001 % sarcosyl
150 mM sodium phosphate (pH 8.5) containing 0.001 % Tween20®
Oligonucleotides: 20 µM
Chemical deactivation of unreacted NHS-esters - 25 mM ethanolamine in
100 mM sodium borate buffer (final pH 8.5)
Incubation (Protein) - 137 mM NaCl, 2.7 mM KCl, 4.3 mM Na2HPO4, 1.4 mM KH2PO4, pH 7.5 with 0.05% Tween20®
Hybridization (DNA) NEXTERION® Hyb (formamide free)
NEXTERION® Oligo Hyb (with formamide)
2x SSC containing 0.1 % SDS and 0.1 % salmon sperm DNA (formamide can be added if required) Add competitor DNA if appropriate

Protocols

Protocol revision

Date: April 2016
Protocol version: 3.0
Revision made: Concretization of storage conditions
Revision reason: Safer storage conditions

Date: April 2013
Protocol version: 2.0
Revision made: New composition of blocking solution
Revision reason: Optimization of blocking and incubation

Date: April 2009
Protocol version: 1.2
Revision made: New telephone and fax numbers
Revision reason: Update contact information
Protocol revision

Date: April 2016
Protocol version: 3.0
Revision made: Concretization of storage conditions
Revision reason: Safer storage conditions

Date: April 2013
Protocol version: 2.0
Revision made: New composition of blocking solution
Revision reason: Optimization of blocking and incubation

Date: April 2009
Protocol version: 1.2
Revision made: New telephone and fax numbers
Revision reason: Update contact information

Date: August 2008
Protocol version: 1.1
Revision made: Changed Tween® concentration (incubation step), pH of blocking solution
Revision reason: Optimization of blocking and incubation



Material Safety Datasheets

Trade Fairs & Events
21.
March
Trade fair Workshop "Microarrays: Basics, Production and Processing", ETH Zurich / University of Zurich, Switzerland, 21.03 - 22.03.2017