BioMicroCenter:BioAnalyzer INFO: Difference between revisions
(→RNA) |
(→DNA) |
||
| (16 intermediate revisions by 2 users not shown) | |||
| Line 1: | Line 1: | ||
{{BioMicroCenter}} | {{BioMicroCenter}} | ||
The Agilent BioAnalyzer 2100 uses nanofluidics to provide quantitative information about biological samples using very small amounts of material. | 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. | ||
==DNA== | ==DNA== | ||
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 preformed relative to the sizing standards. Only 1ul of sample is required. | 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 preformed relative to the sizing standards. Only 1ul of sample is required. The high sensitivity assay is preferred, and samples are diluted to within range unless otherwise requested. | ||
[http://www.chem.agilent.com/en-us/products/instruments/lab-on-a-chip/2100bioanalyzer/dnasolutions/pages/default.aspx DNA] - [http://www.chem.agilent.com/en-us/products/instruments/lab-on-a-chip/pages/gp1565.aspx How it works] ''(Agilent)'' | [http://www.chem.agilent.com/en-us/products/instruments/lab-on-a-chip/2100bioanalyzer/dnasolutions/pages/default.aspx DNA] - [http://www.chem.agilent.com/en-us/products/instruments/lab-on-a-chip/pages/gp1565.aspx How it works] ''(Agilent)'' | ||
| Line 18: | Line 18: | ||
==RNA== | ==RNA== | ||
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 preformed relative to the sizing standards. | 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 preformed relative to the sizing standards. The BioAnalyzer software can generate the [[BioMicroCenter:RIN|RNA Integrity Number (RIN)]], calculate ribosomal ratios of total RNA samples, and detect ribosomal RNA contamination in mRNA. | ||
[http://www.chem.agilent.com/en-us/products/instruments/lab-on-a-chip/2100bioanalyzer/smallrnaassay/pages/default.aspx RNA] - [http://www.chem.agilent.com/en-us/products/instruments/lab-on-a-chip/2100bioanalyzer/rnasolutions/pages/gp38725.aspx Analysis Specifications] ''(Agilent)'' | [http://www.chem.agilent.com/en-us/products/instruments/lab-on-a-chip/2100bioanalyzer/smallrnaassay/pages/default.aspx RNA] - [http://www.chem.agilent.com/en-us/products/instruments/lab-on-a-chip/2100bioanalyzer/rnasolutions/pages/gp38725.aspx Analysis Specifications] ''(Agilent)'' | ||
RNA arrays are segregated by the concentration of RNA used and the type of RNAs being analyzed | 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 | * small RNA: 0.05-2 ng/ul miRNAs | ||
== PROTEIN == | == PROTEIN == | ||
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 | 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 [[BioMicroCenter:High Sensitivity Protein Optimization|optimizing the High Sensitivity assay]]. | ||
Information from Agilent: [http://www.chem.agilent.com/en-us/products/instruments/lab-on-a-chip/2100bioanalyzer/proteinkits/pages/default.aspx Protein] - [http://www.chem.agilent.com/en-us/products/instruments/lab-on-a-chip/pages/gp1566.aspx How it works] ''(Agilent)'' | Information from Agilent: [http://www.chem.agilent.com/en-us/products/instruments/lab-on-a-chip/2100bioanalyzer/proteinkits/pages/default.aspx Protein] - [http://www.chem.agilent.com/en-us/products/instruments/lab-on-a-chip/pages/gp1566.aspx How it works] ''(Agilent)'' | ||
Protein arrays Protein Weight Sensitivity | |||
* Protein 80 5kD- 80kD 60ng/ul | |||
* Protein 230 14kD-230kD 30ng/ul | |||
* High Sensitivity 10kD-250kD 0.3ng/ul | |||
== FLOW CYTOMETRY == | == FLOW CYTOMETRY == | ||
Revision as of 13:19, 11 April 2011
| HOME -- | SEQUENCING -- | LIBRARY PREP -- | HIGH-THROUGHPUT -- | COMPUTING -- | OTHER TECHNOLOGY |
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.
DNA
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 preformed relative to the sizing standards. Only 1ul of sample is required. The high sensitivity assay is preferred, and samples are diluted to within range unless otherwise requested.
DNA - How it works (Agilent)
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 both the DNA 1000 and High Sensitivity DNA kits in stock, if you think you require a different kit please let us know in advance.
RNA
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 preformed 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 - Analysis Specifications (Agilent)
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
PROTEIN
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.
Information from Agilent: Protein - How it works (Agilent)
Protein arrays Protein Weight Sensitivity
- Protein 80 5kD- 80kD 60ng/ul
- Protein 230 14kD-230kD 30ng/ul
- High Sensitivity 10kD-250kD 0.3ng/ul
FLOW CYTOMETRY
In addition to quantification of Nucleic Acid, the BioAnalyzer is capable of preforming simple two color flow cytometry assays. In brief, six samples each 10 µl with 20,000 prestained cells are loaded onto the chip and the fluorescence intensities in two channels for about 750 single cells per sample are measured within 25 minutes.
Information from Agilent: Cells - How it works (Agilent)
