Sample Amplification & Labeling
Sample Amplification & Labeling
Jump to:
Featured Research Using This Product – Genomic DNA Labeling
Featured Research Using This Product – Gene Expression Labeling
Additional Resources
Product Details
Important anouncement for Genomic DNA Enzymatic Labeling Kit
Important information regarding LRILAK Kits
Important product announcement for Genomic DNA Enzymatic Labeling.
See Product Details Section for more information.
Agilent's reagents for sample labeling integrate with our comprehensive DNA microarray platform, offering simple and patented procedures to label genomic DNA for analysis with Oligo aCGH microarrays.
Agilent now offers two genomic DNA Labeling kits:
Genomic DNA Enzymatic Labeling Kit
Genomic DNA ULS Labeling Kit
The Genomic DNA Enzymatic Labeling Kit utilizes an enzymatic methodology of random primers and Klenow to differentially label genomic DNA samples with fluorescently labeled nucleotides. The Genomic DNA ULS Labeling Kit utilizes Kreatech’s Universal Linkage System (ULS™) to chemically bind the label to genomic DNA at the N7 position of guanine. A comparison of the protocols is illustrated in Figure 1. While the performance of both labeling methods is equivalent (Figure 2), the table below outlines the differences between the two kits:
Figure 1. ULS labeling versus traditional enzymatic labeling protocol.
Comparison of the new ULS labeling protocol (on right) versus the traditional enzymatic labeling protocol (on left).
Figure 2. Comparison of Enzymatic and ULS Labeling methods using frozen colon carcinoma.
CGH Analytics v3.4 views of fresh frozen colon carcinoma labeled with either Genomic DNA Enzymatic Labeling kit (left) or Genomic DNA ULS Labeling kit (right). Log2 ratio values for all oligonucleotide probes are plotted as a function of their chromosomal position. The same aberration calls and DLRSD values were observed between the two labeling methods.
The following section describes the utility of the Genomic DNA ULS Labeling Kit with FFPE samples.
FFPE samples are challenging to perform aCGH with because they are often degraded and cross-linked. Thus labeling by traditional enzymatic procedures often results in poor quality aCGH data. By utilizing the Genomic DNA ULS Labeling kit, FFPE samples can be used for aCGH analysis. Using this method we detected and identified identical aberrations in matched FFPE and fresh frozen tissue from the same sample (see Figure 3). We also tested 1-, 4-, and 10-year-old breast cancer FFPE blocks for the well-documented amplification of the human epithelial receptor 2 (HER-2) oncogene and found it was readily identifiable in all three samples (Figure 4).
Figure 3. Chromosomal aberration analysis comparing fresh and frozen FFPE samples from the same tumor.
CGH Analytics v3.4 chromosome views of Chromosome 9 and Chromosome 18 analysis of fresh frozen (blue) and FFPE (green) tissue blocks from the identical tumor. Left panel: Infiltrating ductal breast carcinoma. Right panel: Colon adenocarcinoma. Log2 ratio values for all oligonucleotide probes are plotted as a function of their chromosomal position. Each point represents a single probe with a 1 MB moving average (thin wavy colored vertical lines.) Aberration calls were identified by the ADM-1 algorithm (thick straight vertical colored lines and colored shaded areas).
Figure 4. DNA aberrations in 1-, 4-, and 10-year-old FFPE breast tumor samples.
Detection of Chromosome 17 aberrations in 1- (green), 4- (red) and 10-year-old (blue) infiltrating ductal carcinoma samples showing the common human epithelial receptor 2 (HER-2) gene amplification. Log2 ratio values for all oligonucleotide probes are plotted as a function of their chromosomal position. Each point represents a single probe, with a 1 MB moving average (thin wavy colored vertical lines) on the chromosome panels and no moving average on the zoom-in (3.71 MB) panels. Aberration calls identified by the ADM-1 algorithm (thick straight vertical colored lines and colored shaded areas) are shown. Black arrows show location of HER-2 gene.
Reference: Copy Number Analysis of Archival FFPE Tumor Samples by Oligo Array CGH.
Agilent Publication Number 5989-7120EN.
More
Important product announcement for LRILAK Kits.
See Product Details Section for more information.
Agilent's reagents for sample amplification and labeling integrate with our comprehensive DNA microarray platform, offering simple and patented procedures for one- and two-color labeling of genomic DNA (compatible with Oligo aCGH microarrays) or RNA/cDNA (compatible with Gene Expression microarrays and other downstream analyses). Beginning with as little as 50 ng of total RNA, our highly efficient kits produce up to 2.0 mg of cyanine-labeled cRNA (anti-sense) or cDNA (sense) in only 6 hours, rather than overnight, and require only a single amplification.
Sample limitation is one of common issues in performing DNA microarray experiments, with respect to both isolation and RNA yield from limited tissues and cells. To address the problems specifically associated with insufficient concentrations of labeled cRNA or cDNA, Agilent offers the Low RNA Input Linear Amplification Kit, based on T7 linear amplification and requiring only one round of linear amplification for integration with either oligo or cDNA microarrays.
Figure 1. Generating micrograms of labeled cRNA from nanograms of total RNA after a single amplification.
Using the Low RNA Input Kit, amplified and labeled cyanine-3 or cyanine-5 cRNA was generated from a range of 25 ng to 5,000 ng total RNA from established human cervical carcinomal cell line HeLa cells (A) and human spleen tissue (B), respectively. cRNA yields were determined by UV spectrophotometry by 3 separate users performing triplicates. The HeLa tissue cell line is a rich mRNA source while spleen is a poor source, most likely accounting for the difference in overall yield.
Figure 2. Sample microarray data using total RNA from HeLa and human spleen.
500 ng of cyanine-3-labelled HeLa or cyanine-5-labelled spleen was generated from 25 ng of HeLa or spleen total RNA respectively using the Low RNA Input Kit. They were mixed, and hybridized to an Agilent Human 1A Oligo Microarray using the Agilent In Situ Hybridization Kit Plus. Array image generated by the Agilent dual-laser DNA microarray scanner with SureScan technology (A). Log intensity plot from Rosetta Resolver® analysis (B). Blue "+" indicates there is no differential gene expression between HeLa and spleen, while red “+” represents upregulated gene expression, and green “+” represents downregulated gene expression, in spleen compared to HeLa (p <= 0.01).
Reference: Efficient Method for Isolation of High Quality Concentrated Cellular RNA with Extremely Low Levels of Genomic DNA Contamination.
Agilent Publication Number 5989-0322EN.
More
Listen to Agilent researchers present current product information.
View available eSeminars
PRODUCT UPDATE: Agilent introduced the Genomic DNA Enzymatic Labeling Kits on August 1, 2008. this kit is a direct replacement of the Genomic DNA Labeling Kit PLUS. To learn more, download the latest tech note: A Comparison of the Genomic DNA Enzymatic Labeling Kit and the Genomic DNA Labeling PLUS Kit.
Genomic DNA ULS Labeling Kit - Chemical labeling of genomic DNA
For detailed specifications and ordering information visit agilent.com
Genomic DNA Purification Module - The purification module is modified to be compatible with Agilent Oligo aCGH microarrays
For detailed specifications and ordering information visit agilent.com
Genomic DNA Enzymatic Labeling Kit - NEW: validated and optimized for streamlined, enhanced performance in Agilent's Oligonucleotide Array-Based CGH workflow
For detailed specifications and ordering information visit agilent.com
Quick Amp Labeling Kit, No Dye - NEW: improved reagents used to amplify total RNA for use as targets in gene expression studies
For detailed specifications and ordering information visit agilent.com
Quick Amp Labeling Kit, One-Color (Cy3) – NEW: improved reagents used to amplify and label total RNA for use as targets in one-color gene expression studies
For detailed specifications and ordering information visit agilent.com
Quick Amp Labeling Kit, Two-Color (Cy3 and Cy5) – NEW: improved reagents used to amplify and label total RNA for use as targets in two-color gene expression studies
For detailed specifications and ordering information visit agilent.com
PRODUCT UPDATE: LRILAK Kits have been replaced by Quick Amp Labeling Kits. If you require a large number of LRILAK kits to finish a study, please contact your local sales representative. To learn more, download the the latest tech note: A Comparison of Agilent Quick Amp and Low RNA Input Linear Amplification Kits.
miRNA Labeling Reagent and Hybridization Kit: provides a novel Cyanine 3-nucleotide along with array hybridization reagents to selectively label and hybridize mature miRNAs for Agilent's miRNA Microarray System
For detailed specifications and ordering information visit agilent.com
For research use only and not for use in diagnostic procedures.