Service is provided for projects utilizing Affymetrix GeneChips or glass slide microarrays. Both types of projects start with at least one consultation with Dr. Tapan Ganguly to review experimental design and the various assay options and requirements. Total RNA extraction and purification is offered for clients who do not have molecular biology resources.

Affymetrix services include:

  • Target preparation cDNA synthesis, in vitro transcription, purification, fragmentation, quality control. Double amplification for projects supplying 50ng, 3µg total RNA.
  • GeneChip hybridization, washing, staining, scanning.
  • Data formatting and save to disk, chip-to-chip pairwise analyses, quality control report.

Glass slide services include:

  • E. coli colony picking, clone set propagation, plasmid or BAC extraction.
  • PCR set-up, thermocycling, PCR clean-up.
  • Print plate preparation for oligomer or PCR probe sets.
  • Glass slide array design and printing.
  • Automated slide hybridization and washing.
  • Dual channel fluorescence scanning.
  • Consultation on experimental design, target labeling and array processing.

    GLASS SLIDE MICROARRAYS

    • Mouse embryo libraries 15,000 PCR products from the NIA cDNA collection
    • Mouse 15,000 long oligomers, MWG set
    • Human 19,000 long oligomers, Compugen set
    • Human BAC 4000 PCR products from mapped BACs for CGH, Weber et al.
    • Yeast 6400 long oligomers, Qiagen
    • Pig 11,000 long oligomers, Qiagen

    SELF-SERVICE EQUIPMENT

    • Tecan HS400 Hyb Station automated glass slide processing including pre-hyb, hybridization, washing, drying. Capacity 2 x 12 slides. Hyb volume about 100ul.
    • Axon GenePix 4000B scanner dual laser, two-channel detection of fluorescence on glass microscope slides. Excitation 532nm and 635nm. Detection 557-592nm and 650-690nm. 5 micron resolution.
    • Genicon RLS scanner resonance light scattering detection of gold or silver microspheres. Highly sensitive detection of antibody labeled targets on glass slide microarrays.

    Massively Parallel Sequencing

    Equipment description

    Quantitative PCR and Molecular Diagnostics

    Gene Expression Assay
    SNP Genotyping
    Investigator Specific Assays

    Apo E Genotyping Apolipoproteins are proteins that bind and transport lipids in blood and tissues. The complex of protein and lipid is a lipoprotein. Several major apolipoproteins have been described which differ in their structure, physicochemical behavior, function and distribution. Apolipoprotein E (ApoE) binds to the LDL receptor and serves as a ligand for receptor-mediated endocytosis.

    Genetic defects of apolipoproteins cause various abnormalities in lipid metabolism and increased susceptibility to heart disease. The ApoE gene spans 3.7 kb including 4 exons and is located on chromosome 19. ApoE allelic frequencies vary among different populations. ApoE is a susceptibility gene that contributes to the risk of developing Alzheimer's disease (AD) after age 65 in demented individuals. Three isoforms of ApoE exist: E2, E3, and E4. A demented patient harboring two ApoE4 alleles has a 91% chance of developing AD by age 80, whereas a demented patient harboring one E4 allele has a 47% chance of developing the disease by age 80. ApoE genotyping is useful both as an adjunct to help confirm a suspected clinical diagnosis of AD, as well as a tool to help differentiate AD from atypical presentations of dementia.

    Chimerism Analysis for Donor Lymphocyte Infusion Adoptive immunotherapy, using the infusion of allogeneic immunocompetent leukocytes, has proven to be a promising strategy for the treatment of various hematologic malignancies. Chimerism analysis after such therapy can be predictive of treatment failure and/or relapse. This is a reliable method to detect low levels of mixed chimerism. However, relapse remains a major cause of treatment failure. The relapse risk is associated with remaining recipient cells in the marrow and circulation rather than total removal of the recipient's own cells and replacement by donor cells. This test can be used to initially document engraftment of donor cells and to detect residual or recurrent recipient cells as early as possible before relapse so that appropriate therapy can be instituted.

    Gene Expression Assay

    Gene expression changes provide specific physiological signals that are associated with early cellular responses. The strategy to identify, validate and optimize gene targets and gene analysis systems for prompt examination of gene expression biomarkers will be useful to identify diseased condition.

    Identity Testing The genes make up only a tiny fraction of the DNA. The rest, the great bulk - about 97 percent - has no known function. It is sometimes referred to as "junk DNA." Nevertheless, these nongenic regions show the same genetic variability that genes do, in fact usually more. These differences are not overt, but can be detected by laboratory tests. Regions of DNA that are used for forensic analysis are usually not genes, but rather are located in those parts of the chromosomes without known functions, or if part of a gene, not in the part that produces a detectable effect. Nevertheless, the words commonly used for describing genes (e.g., allele, homozygous, polymorphic) are carried over to DNA regions used for identification. It is customary to call the genotype for the group of loci involved in an identity analysis profile.

    SNP Genotyping Single nucleotide polymorphisms (SNPs) are common DNA sequence variations among individuals. On average, SNPs occur in the human population more than 1 percent of the time. Because only about 3 to 5 percent of a person's DNA sequence codes for the production of proteins, most SNPs are found outside of "coding sequences" and they serve as biological markers for pinpointing a disease on the human genome map. SNPs found within a coding sequence are of particular interest to researchers because they are more likely to alter the biological function of a protein. They promise to significantly advance our ability to understand and treat human disease and have become central to the advancement of molecular genetics.

    Using SNPs to study the genetics of drug response will help in the creation of "personalized" medicine, as SNPs may also be associated with the absorbance and clearance of therapeutic agents.

    T Cell Receptor V-β Analysis (CDR3 Spectratyping) This screening can be used for a qualitative assessment of clonal diversity among peripheral blood T cells, T-cell lines and clones emerging from bulk and limiting dilution cultures. It can also be used as a tool for assessing the clonal composition of T-cell populations.

    Twin Zygosity DNA Test This is also identity testing. Twins are either dizygotic (commonly referred to as fraternal) or monozygotic (identical). Dizygotic twins are the result of two different eggs being fertilized by two separate sperm and resulting in two completely distinct and separate pregnancies in the womb at the same time. Monozygotic twins are the result of one egg fertilized by a single sperm. Twin Zygosity refers to the genetic relationship between twin siblings. The Twin Zygosity DNA Test is used to establish DNA profiles for the twins being tested. If the profiles are different, then the twins are fraternal, and if the profiles are identical then a probability of being identical twins is calculated.