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| Description |
Gene expression profiling
from 1 – 10,000 cells
Cells sorted with MACS® Technology, laser capture microdissected cells, cells sorted by flow cytometry, tissue biopsies and primary cells can be analyzed using the SuperAmp™ Service: RNA is sensitively isolated and amplified from even a few cells.
Send your sample...
The SuperAmp™ Preparation Kit enables a secure, convenient sample lysis and storage. Subsequently, samples are sent to Miltenyi Biotec on dry ice, where amplification of RNA is performed and documented.
...and receive ready-to-publish data
The service team processes Agilent Whole Genome Microarrays as well as topic-defined or custom PIQOR™ Microarrays, and provides in-depth bioinformatic analysis. An extensive report and interpreted results are received within an average of three weeks. |
| Applications |
Unique RNA amplification protocol
The SuperAmp Service uses global PCR for the amplification of mRNA-derived cDNA. mRNA is sensitively isolated via magnetic bead technology. Linear amplification of tiny amounts of RNA is achieved and PCR bias avoided by:
- Single-primer global PCR with uniform annealing conditions for all transcripts
- Consistent length of generated cDNA fragments
Detection of differentially expressed genes down to a single cell
mRNA was extracted from 1; 10; 100; 1,000; 10,000; and 107 Jurkat and Raji cells, respectively. After reverse transcription, cDNA was globally amplified, then labeled and hybridized to human PIQOR Immunology Microarrays.
The cluster analysis in figure 1 shows virtually no difference of expression profiles between the different amounts of starting material. Differentially expressed genes are reproducibly detected starting with 10,000 cells down to one single cell – also compared to 107 cells without amplification of mRNA.
Expression profiling analysis of NK cell subsets
Four natural killer cell subsets from two donors have been purified using MACS Technology: CD56+CD16–; CD56dimCD16+ cells; CD56+CD8+ cells; CD56+CD8– cells. mRNA isolation and amplification from 1,500 cells was performed according to the SuperAmp™ Protocol, followed by microarray analysis based on human PIQOR™ Immunology Microarrays. The cluster analysis in figure 3 indicates that CD56+CD16– cells show the greatest difference to the other cell subsets. The mainly differentially expressed genes are highlighted in panel 1 for downregulated genes and in panel 2 for upregulated genes. |
| Logistics |
Order...
The SuperAmp Service is ordered in combination with the PIQOR Microarray Service Plus Amplification or the Agilent Whole Genome Microarray Service.
Please fill in the "Service Order Confirmation" form, available as a download at www.miltenyibiotec.com.
...receive SuperAmp Preparation Kit...
The SuperAmp Preparation Kit is sent to you for the lysis and storage of your valuable samples. Please use only the Preparation Kit for sample lysis and storage.
...and ship samples
The samples are shipped on dry ice to Miltenyi Biotec where SuperAmp Service is performed followed by the Microarray Service. |
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| Figure 1 |
Cluster Analysis
Jurkat versus Raji cells with different amounts of starting material. |
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Gene expression:
red = increase;
green = decrease;
black = no alteration;
gray = no signal detection.
x-axes: array experiments;
y-axes: gene names. |
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| Figure 2 |
| Pearson correlation coeffcient of repeated microarray experiments shows that the results are technically and biologically reproducible down to 100 cells. Below 100 cells, the reproducibility is reduced due to the enhanced influence of individual cells. |
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| Figure 3 |
Cluster Analysis
Four NK cell subsets from two donors. |
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Gene expression:
red = increase;
green = decrease;
black = no alteration;
gray = no signal detection.
x-axes: array experiments;
y-axes: gene names. |
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