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The Agilent BioAnalyzer 2100 uses nanofluidics to provide quantitative information about biological samples using very small amounts of material. An application compendium for DNA, RNA, Protein and Cell analysis can be found in the BioMicro Center.
The BioAnalyzer analyzes DNA using the same principles as slab agarose gels. A very small tube of gel is created and a fraction of the DNA loaded, along with a low and high size marker, and are placed at one end of the chip. Then, using a voltage gradient, the sample is run down the length of the chip. As the DNA passes a laser, it is detected. Subsequent samples are run down the same lane of the chip. Quantification is performed relative to the sizing standards. Only 1 uL of sample is required. The high sensitivity assay is preferred, and samples are diluted to within range unless otherwise requested.
There are four different options for DNA arrays:
- DNA 1000 Kit: 25bp - 1,000bp, 0.1-50ng/uL
- DNA 7500 Kit: 100bp - 7,500bp, 0.1-50ng/uL
- DNA 12000 Kit: 100bp - 12,000bp, 0.1-50ng/ul
- High Sensitivity DNA kit: 50bp - 7,000bp, 5-500pg/ul
The BioMicro Center keeps only the High Sensitivity DNA kit in stock.
The BioAnalyzer analyzes RNA using the same principles as denaturing slab agarose or polyacrylamide gels. A very small tube of gel is created and a fraction of the RNA loaded, along with a low marker, is placed at one end of the chip. Then, using a voltage gradient, the sample is run down the length of the chip. As the RNA passes a laser, it is detected. Subsequent samples are run down the same lane of the chip. Quantification is performed relative to the sizing standards. The BioAnalyzer software can generate the RNA Integrity Number (RIN), calculate ribosomal ratios of total RNA samples, and detect ribosomal RNA contamination in mRNA.
RNA arrays are segregated by the concentration of RNA used and the type of RNAs being analyzed
- RNA Nano: 25-500 ng/ul total RNA
- RNA Pico: 0.05-5 ng/ul total RNA
- small RNA: 0.05-2 ng/ul miRNAs
The same principles for RNA and DNA apply for protein. Here, the model is SDS-PAGE with samples denatured before loading and run in reducing (+DTT) or non-reducing conditions. The standard and high sensitivity assays are moderately different from each other. Standard assays work like DNA and RNA in that the sample is added to a dye and run together in the lane. With the high-sensitivity system, the buffers are exchanged by standard chromatography to tune the pH and remove contaminating chemicals before being labeled. The labeled proteins then are run on the chip - or can be used for additional purification steps before running. BMC is currently optimizing the High Sensitivity assay.
Protein arrays Protein Weight Sensitivity
- Protein 80 5kD- 80kD 60ng/ul
- Protein 230 14kD-230kD 30ng/ul
- High Sensitivity 10kD-250kD 0.3ng/ul
In addition to quantification of Nucleic Acid, the BioAnalyzer is capable of performing simple two color flow cytometry assays. In brief, six samples each at concentration of 20,000 prestained cells with a minimum of 10 µL are loaded onto the chip and the fluorescence intensities in two channels for about 750 single cells per sample are measured within 25 minutes.