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| μMACS™ One-step cDNA Labeling Kit |
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Three steps in one column – mRNA isolation, cDNA labeling, and cDNA purification with MACS® Technology.
95 minutes from sample to microarray hybridization – with fluorescently or radioactively labeled first-strand cDNA.
Sensitivity is the key – the extremely small MicroBeads and the in-column procedure significantly minimize loss of sample for sensitive results.
Reproducible cDNA labeling – the convenient MACS Technology and ready-to-use reagents reduce operator variance, hands on time, and sample peparation variability.
Innovation in cDNA labeling
Gene expression analysis with microarrays generally starts with RNA isolation, followed by cDNA labeling and purification of the hybridization probe. Accuracy of each of these steps is very important for sensitive and reproducible results. The magnetic isolation and in-column processing with MACS Technology not only combines these three steps in one column, but is also designed for highest quality of results. The isolation of messenger RNA, the minimized loss of material during processing and purification, as well as the convenient handling increase sensitivity and reproducibility.
MACS® Technology for gene expression profiling
Extremely small (diameter 50 nm), paramagnetic Oligo(dT) MicroBeads instantly label polyA+ mRNA for purification directly from cells or tissues as well as from total RNA (fig. 1). In the strong magnetic field of a thermoMACS™ Separator, these labeled mRNA molecules are retained while impurities are washed away. Now the ready-to-use enzyme mix with the label of choice – radioactive or fluorescent – is added, and the thermoMACS Separator is switched on for a 60-minute incubation at 42 °C. Final washing and an RNase H step remove reaction components, unincorporated nucleotides, and residual RNA. The elution yields pure, labeled first-strand cDNA that can directly be used in a microarray hybridization.
Consistent cDNA labeling for microarray experiments
In repeated hybridization experiments with PIQOR™ Immunology Microarrays, sense, the successful combination of μMACS™ mRNA isolation with in-column cDNA labeling could be visualized. Figure 2 shows a scatter plot of a self-versus-self hybridization of Cy3- and Cy5-labeled mouse liver probes. The high regression coefficient of 0.994 demonstrates that both labeled cDNAs give nearly identical signals on the microarray. |
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| Figure 1 |
| μMACS One-step cDNA Labeling |
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| Figure 2 |
| Scatter plot, self-self hybridization of PIQOR™ Immunology Microarray, sense, with probes generated from 10 mg mouse liver and labeled either with Cy3-dCTP or Cy5-dCTP. The high regression coefficient of 0.994 demonstrates that both labeled cDNAs give nearly identical signals on the microarray. |
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| Figure 3 |
| Pearson correlation-coefficient visualizing reproducibility and reliability of one-step cDNA labeling. 10 μg and 100 μg total RNA derived from Molt-4 cells were labeled with Cy3 versus 10 μg and 100 μg total RNA isolated from Raji cells labeled with Cy5. Corresponding samples were hybridized to PIQOR™ Immunology Microarrays, sense, using the a-Hyb™ Hybridization Station. |
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| Products |
| μMACS One-step cDNA Labeling Kit |
- for 20 reactions
Components
- 1 mL Oligo (dT) MicroBeads
- 40 mL Lysis/Binding Buffer
- 20 mL Wash Buffer
- 20× Lyophilized cDNA Labeling Mix
- 20× Lyophilized RNase H
- 2× 0.5 mL Resuspension Buffer
- 2× 15 mL Equilibration/Wash Buffer
- 0.5 mL cDNA Release Solution
- 5 mL cDNA Elution Buffer
- 100 μL μMACS Sealing Solution
- 20 μ Columns
- 20 LysateClear Columns
Download datasheet
130-092-443
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| μMACS One-step cDNA Labeling Starting Kit |
- for 20 reactions
Components
- 1 μMACS One-step cDNA Labeling Kit (20 reactions)
- 1 thermoMACS™ Separator
- 1 MACS MultiStand
130-092-521
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