CUSTOM MICROARRAY PRINTINGWe custom design arrays according to client requirements and print them on membranes or slides of any particular chemistry. We also provide consultation in preparation of sample plates and printing programs before undertaking any printing project. The consultation ranges from recommendation on the type of micro-well plates to use for array samples to be printed, location of such samples in the plate so they appear at the desired places on the array, and where the desired replicates and control samples should appear on the arrays. Printed arrays are shipped along with the corresponding gal file to clients via courier service. Alternatively, the printed arrays are processed for hybridization at KamTek and the data files are sent to the clients.
Arrays are printed with ChipwriterTM Pro Technology based on super-precision robotics, neuro-fuzzy control and proprietary PC/104 motion controller operated by software developed by Virtek Biotechnology. The subassemblies are carefully integrated by using structural design methods for zero deflection, specialized alignment techniques, and sub-micron level instrumentation to achieve the precision and repeatability required to dispense various patterns of arrays uniformly. The robotic cell is used for high throughput spotting on glass slides and membranes
SALIENT FEATURES OF MICROARRAY PRINTING SERVICES
- Samples are printed with Well Established Robotics and Microfluidic Systems in HEPA Filtered Clean Air and Humidity Controlled Chambers.
- A variety of substrates with specific surface chemistries from different manufacturers or clients can be used for printing.The substrates could be glass slides, plastic slides or membranes. Appropriate printing substrate could be selected by the client.
- Printing on ultra clean, ultra flat slides from Telechem and Corning (UltraGAP) gives best results for DNA arrays in our hands. In most cases, these slides do not need blocking before hybridization.
- Quill-Pins (Stealth Pins) can be used for printing on glass/plastic slides or membranes. Solid-Pins are used for Printing on membranes.
- The microarray spot size can be varied from 100micron to 150micron and the center-to-center distance between spots can be adjusted between 200-450microns.
- We can print high-density arrays up to 15,000 spots arranged in 4-48 blocks (grids) per slide. Spots can be printed in replicates placed either next to each other or in separate grids. We work with the client to design their array pattern.
- Platen can accommodate up to 100 slides per printing batch.
- Immobilization of DNA printed on glass slides is achieved either by UV or heat as desired by the client.
- An appropriate gal-file or text file accepted by a variety of scanners is provided with the final product.
ARRAY MATERIAL PREPARATIONArrays are printed with PCR product DNA, oligo-nucleotides (oligos), proteins, lectins, glyco-proteins, and a variety of sugar molecules.
There are two major types of DNA arrays. Arrays prepared by in situ synthesis of oligos or by printing or spotting of DNA preparations which could be either PCR products or synthetic oligos. Whereas In situ synthesis of oligos has its own advantages, it does not allow custom design of microarrays, or independent evaluation and purification of oligos before their attachment to microarray. PCR product or pre-synthesized oligo (50-70mers) preparations avoid these limitations and in addition provide a larger region of inquiry.
PCR products: Clients can supply their own purified DNA preparations with authenticity of each preparation having been fully established by them. Alternatively, we provide a service for PCR product sample preparation from bacterial clones. This includes amplification of Plasmid DNA from bacterial clones, purification of amplified DNA preparation; determine the authenticity of the amplified material, quantitation and reconstitution of purified DNA at the desired concentrations of 200-300ng/ul for printing.
Oligos: Oligo arrays are gaining popularity due to availability of gene sequences for increasing number of species, and wide spread availability of high throughput synthesis of long oligos (50-70mers) at a relatively low cost, high reproducibility and fast turn around time. The latter includes quality check of each oligo during synthesis, capillary electrophoresis or mass spectroscopy of each oligo after synthesis, and well to well normalization of each preparation. Clients can provide sequences of the oligos to be printed or they can provide already synthesized 50-70mer oligos reconstituted at the required concentrations in printing buffers of their choice and distributed in 384 well plates.
Most proteins are printed in PBS or in PBS with 20-50% glycerol for antibodies. The printing concentrations range from 0.1-1ug/ul. There are glass slides available from different manufacturers with different chemistries, and therefore require different printing and processing protocols. We work with our clients in the selection of substrates, appropriate printing buffers, and for design of arrays. We can also print protein arrays on membranes and membrane coated slides. Samples should be supplied frozen as specified for PCR products.
A variety of molecules containing carbohydrate moieties, such as oligosaccharides, glycoproteins, glycolipids, glycoconjugates and glycomimetics can be printed on a variety of solid substrates. These microarrays could be prepared on some glass substrates simply by adsorption or by covalent binding procedures. The latter may require further derivatization of solid substrates and/or carbohydrate molecules themselves. We can help our clients to establish optimized procedures for their varied experimental requirements
PREPARATION OF SAMPLES AND SHIPPING REQUIREMENTS
KTI has worked out optimal concentrations of samples to have probe excess and uniform spot morphology and recommend these concentrations for printing. Sample preparations can be supplied in either 384 well (Genetix Cat. # 7020) or 96 well polypropylene plates with loosely fitted lids (Cat. # 7021). Each plate should be sealed with a sealing mat (Genetix cat. # 5150). Samples should be shipped frozen to the KamTek facility. We also require an excel file containing information on sample location and identification to generate gal file for scanning and analysis purposes.
Alternatively, clients can also provide bacterial clones to prepare PCR product DNA at our facility or provide sequences of oligos, which can be synthesized and reconstituted at our facility.
QUALITYWe follow Standard Operating Procedures including pre-established quality control and quality assurance protocols. Pictures of microarrays are included for clients reference. To monitor the quality of printing for QC purposes, we recommend including standard PCR product DNA samples or oligos for house keeping genes along with experimental samples. To monitor quality of total microarray experiments, we recommend printing of samples in triplicates.
TURN AROUND TIME
- Two to three days for printing services with ready to go preparations of DNA, Oligomer, protein, glyco-protein, lectin or polysaccharide molecules in buffers of choice and at recommended concentrations.
- Four to six weeks for preparation, purification, QC analysis, quantitation and dispensing desired concentrations of PCR products in 384-well plates followed by DNA microarray printing, and
- One week for oligo synthesis, QC analysis, quantitation and dispensing desired concentrations of oligos in 384-well plates followed by Oligo- microarray printing.