Agilent Technologies 2100 Bioanalyzer, Illumina Beadstation, Tecan robotics, Tecan 400 Pro hybridization station, Agilent 4200 Scanner and assorted small instrumentation. MicroArray samples are run on an Agilent 2100 biochip for quality control and must pass QC to be used. A Nanodrop instrument reading is also taken prior to the start of any processing.
While the Microarray core can process arrays commercially purchased from Agilent, Nugene, the most powerful platform are bead based arrays from Illumina. These arrays are based on 3 um silica beads: (Insert photo).
Each array has about 1×106 beads with each transcript replicated ~30 times. The results give extraordinary statistics. –(insert photo)
High density SNP profiling (Illumina) and microRNA arrays (Exiqon 2 color ) are also available. Preliminary data analysis assistance is always provided with results provided in an Excel workbook that includes: All genes on the array, all genes expressed in each sample with p ‹0.01. all genes differentially expressed (typically p ‹0.001) between groups. This data can easily be exported for further analysis. The core supplies a computer and access into Ingenuity, a pathway software package, for a small user fee.
For more information please make an appointment to speak with Dr. David Loose at 713-500-7440.
RNA & DNA Quantification
Total RNA should be clean as possible. A column clean up method is highly recommended. Amounts of RNA needed vary depending on the platform and study being done. For Illumina expression arrays 10 µl of 100 ng/µl is usually required. For any other formats and lower amounts of RNA contact the lab for help in determining amounts needed.
For use with Nucleic Acids real-time qPCR instruments- Applied Biosystems 7900 (384-well) and Roche LC480 (96 or 384-well) are used.
RNA quality must be monitored prior to running real-time qPCR plate screening arrays (Exiqon, Roche, ABI, Qiagen, Lonza, etc). Samples are run on an Agilent 2100 biochip for quality control and must pass QC to be used for the succeeding real-time qPCR plate arrays. A Nanodrop instrument is available for outside users for an annual fee. We also have a Tecan Infinite 200 multimode plate reading spectrofluorometer multi-mode reader for fluorescent determination of DNA or RNA concentrations. Again, an annual user fee applies for this instrument for unlimited use. Prior to measuring miRNAs in serum/plasma quality is determined using a small number of miRNA assays. All real time qPCR assays are setup using robotic liquid handlers, Tecan Genesis 100 (RT setup or additions of DNA samples) a Beckman Biomek3000 (PCR master mix).
Assay modalities (all 384-well plate determinations except where mentioned below):
- Nucleic acid detection and quantification, over 1,000 validated real-time qPCR assays available.
- miRNA screening: Exiqon LNA-based primers, 2 384-well plate arrays for human, mouse and rat utilizing sensitive qPCR detection and GenEx software for data analysis. Focused arrays and custom arrays are also available.
- Validation of miRNA (Exiqon), microarray, siRNA knockdown or compound screening ‘hits’.
- Zygosity testing of mouse litters for endogenous or transgenes.
- Plate screening arrays from SA Biosciences/Qiagen, Lonza StellArrays, Life Technologies and Roche plate arrays for focused pathway or cell physiology determinations (96- or 384-well plates).
- High resolution melt (HRM) analysis of SNPs, mutations, species differentiation, methylation and more. Inquire concerning this new application for real-time PCR.
- Protein melt analysis for small compound/purified protein association screening studies. Can replace a fluorescence polarization assay if the association is strong enough, for less cost per assay.
MSD 2400 CCD camera electrochemiluminescence plate reader is used for MSD quantitative sandwich single or multiplex ELISA assay plates.
Biomek NXp 96-tip head liquid handling robotic workstation with integrated ThermoFisher Cytomat tip and plate store and Bio-tek ELx405 automated plate washer as well as a Biomek3000 are used for sample dispensing and setting up standard curves.
Assays: Any Meso Scale Discovery assay kit. Contact Dr. Loose via phone at (713) 500-7440 or via e-mail at David.S.Loose@uth.tmc.edu for the latest information on kits. We can help with the development of novel assays on the MSD platform using single spot plates and reagents from MSD.
For information on Mass Spectrometry analysis of proteins and insight into protein purification and handling, contact the Clinical and Translational Proteomics Service Center.
We also have an ABI 7700s available to outside users with a flat fee/plate charge. Signup sheets are on the instruments.