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Agilent DNA 7500 Kit Analyzer Quick Start Guide PDF

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Summary of Content for Agilent DNA 7500 Kit Analyzer Quick Start Guide PDF

Agilent DNA 7500 and DNA 12000 Kit Quick Start Guide

Assay Principles Agilent DNA kits contain chips and reagents designed for sizing and analysis of DNA fragments. Each Agilent DNA contains an interconnected set of microchannels that is used for separation of nucleic acid fragments based on their size as they are driven through it electrophoretically. Agilent DNA kits are designed for use with the Agilent 2100 Bioanalyzer instrument only.

Assay Kits Agilent DNA 7500 and DNA 12000 kits are designed for the sizing and quantitation of double-stranded DNA fragments from 100 to 7500 base pairs (DNA 7500) and 100 to 12000 bp (DNA 12000 respectively). These kits can be used to analyze, for example, PCR and RT-PCR products as well as restriction digests. The complete DNA 7500 and DNA 12000 kit guide can be found in the online help of the 2100 Expert Software.

Agilent DNA 7500 Kit (reorder number 5067-1506) and Agilent DNA 12000 Kit (reorder number 5067-1508) DNA Chips Reagents1& Supplies

1 DNA 7500 Reagents (reorder number 5067-1507); DNA 12000 Reagents (reorder number 6067-1509)

25 DNA Chips (yellow) DNA Ladder 1 Electrode Cleaner (green) DNA Markers (2 vials) Syringe Kit (blue) DNA Dye Concentrate 2(1 vial)

2 This product is provided under a license by Life Technologies Corporation to Agilent Technologies. The purchase of this product conveys to the buyer the non-transferable right to use the purchased amount of the product and components of the product only as described in accompanying product literature. The sale of this product is expressly conditioned on the buyer not using the product or its components (1) in manufacturing; (2) to provide a service, information, or data to an unaffiliated third party for payment; (3) for therapeutic, diagnostic or prophylactic purposes; (4) to resell, sell or otherwise transfer this product or its components to any third party, or use for any use other than use in the subfields of research and development, quality control, forensics, environmental analysis, biodefense or food safety testing. For information on purchasing a license to this product for purposes other than described above contact Life Technologies Corporation, Cell Analysis Business Unit, Business Development, 29851 Willow Creek Road, Eugene, OR 97402, Tel: (541) 465-8300. Fax: (541) 335-0354.

1 Syringe (red) DNA Gel Matrix (3 vials) 3 Spin Filters

Agilent Technologies

Agilent DNA 7500 and DNA 12000 Kit Quick Start Guide

Storage Conditions Keep all reagents and reagent mixes refrigerated at 4 C when not in use to avoid poor results caused by

reagent decomposition. Protect dye and dye mixtures from light. Remove light covers only when pipetting. Dye decomposes when

exposed to light.

Equipment Supplied with the Agilent 2100 Bioanalyzer Chip priming station (reorder number 5065-4401) IKA vortex mixer

Additional Material Required (Not Supplied) Pipettes (10 L, 100 L and 1000 L) with compatible tips 0.5 mL microcentrifuge tubes for sample preparation Microcentrifuge

Sample Preparation For accurate determination of DNA concentration, the total amound of DNA in sample must be between

0.550 ng/l. If concentration is excessively high, dilute to 0.550 ng/l. When analyzing restriction digests, add EDTA or heat inactivate the restriction enzyme according to the

manufacturer's instruction. Restriction endonucleases in combination with non-chelated metal ions may degrade internal DNA marker.

Physical Specifications Analytical Specifications

Type Specification Type Agilent DNA 7500 Agilent DNA 12000 Analysis time 30 min Sizing range 100 7500 bp 1000 12000 bp Samples per chip 12 Sizing resolution 100 1000 bp: 5 %

1000 7500 bp: 15 % 100 1000 bp: 5 % 1000 12000 bp: 15 %

Sample volume 1 L Sizing accuracy 10 % 15 % Kit stability 4 months (see box

for storage temperatures)

Sizing reproducability

5 % CV 5 % CV

Kit size 25 chips 12 samples/chip = 300 samples/kit

Quantitation accuracy1

1 Determined using the respective DNA ladder as sample

20 % 25 %

Quantitation reproducibility1

100 1000 bp: 10 % CV 1000 7500 bp: 5 % CV

100 1000 bp: 15 % CV 1000 12000 bp: 10 % CV

Quantitative range1

0.5 50 ng/L 0.5 50 ng/L

Maximum salt concentration in sample

250 mM for KCl, 15 mM for MgCl2, 250 mM NaCl

250 mM for KCl, 15 mM for MgCl2, 250 mM NaCl

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Agilent DNA 7500 and DNA 12000 Kit Quick Start Guide

Setting up the Chip Priming Station 1 Replace the syringe:

a Unscrew the old syringe from the lid of the chip priming station. b Release the old syringe from the clip. Discard the old syringe. c Remove the plastic cap of the new syringe and insert it into the clip. d Slide it into the hole of the luer lock adapter and screw it tightly to the chip

priming station.

2 Adjust the base plate: a Open the chip priming station by pulling the latch. b Using a screwdriver, open the screw at the underside of the base plate. c Lift the base plate and insert it again in position C. Retighten the screw.

3 Adjust the syringe clip: a Release the lever of the clip and slide it up to the top position.

Essential Measurement Practices Handle and store all reagents according to the instructions on the label of the

individual box. Avoid sources of dust or other contaminants. Foreign matter in reagents and

samples or in the wells of the chip will interfere with assay results. Keep all reagents and reagent mixes refrigerated at 4 C when not in use. Allow all reagents and samples to equilibrate to room temperature for 30 min before use. Protect dye and dye mixtures from light. Remove light covers only when pipetting. The dye decomposes

when exposed to light and this reduces the signal intensity. Always insert the pipette tip to the bottom of the well when dispensing the liquid. Placing the pipette at

the edge of the well may lead to poor results.

Use a new syringe and electrode cleaners with each new kit. Use loaded chips within 5 min after preparation. Reagents might evaporate, leading

to poor results. Do not touch the Agilent 2100 Bioanalyzer during analysis and never place it on a

vibrating surface.

Agilent DNA 7500 and DNA 12000 Assay Protocol - Edition April 2007

WARNING Handling DMSO Kit components contain DMSO. Because the dye binds to nucleic acids, it should be treated as a potential mutagen and used with appropriate care. Wear hand and eye protection and follow good laboratory practices when preparing and handling

reagents and samples. Handle the DMSO stock solutions with particular caution as DMSO is known to facilitate the

entry of organic molecules into tissues.

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Agilent DNA 7500 and DNA 12000 Kit Quick Start Guide

Preparing the Gel-Dye Mix 1 Allow DNA dye concentrate (blue ) and DNA gel matrix (red ) to equilibrate to room temperature for

30 min.

2 Vortex DNA dye concentrate (blue ) and add 25 L of the dye to a DNA gel matrix vial (red ).

3 Vortex solution well and spin down. Transfer to spin filter. 4 Centrifuge at 1500 g 20 % for 10 min. Protect solution from light. Store at 4 C.

Loading the Gel-Dye Mix 1 Allow the gel-dye mix to equilibrate to room temperature for 30 min before use. 2 Put a new DNA chip on the chip priming station.

3 Pipette 9 L of gel-dye mix in the well marked . 4 Make sure that the plunger is positioned at 1 mL and then close the chip priming station. 5 Press plunger until it is held by the clip. 6 Wait for exactly 30 s then release clip. 7 Wait for 5 s, then slowly pull back the plunger to the 1 mL position.

8 Open the chip priming station and pipette 9 L of gel-dye mix in the wells marked .

Loading the Marker 1 Pipette 5 L of marker (green ) in all sample and ladder wells. Do not leave any wells

empty.

Loading the Ladder and the Samples

Technical Support Please visit our support web page http://www.agilent.com/genomics/contactus to find information on your local Contact Center.

Further Information Visit the 2100 Bioanalyzer site at http://www.agilent.com/genomics/bioanalyzer. You can find useful information, support and current developments about the products and the technology.

1 Pipette 1 L of DNA ladder (yellow ) in the well marked . 2 In each of the 12 sample wells pipette 1 L of sample (used wells) or 1 L

of de-ionized water (unused wells). 3 Put the chip horizontally in the adapter and vortex for 1 min at the

indicated setting (2400 rpm). 4 Run the chip in the Agilent 2100 Bioanalyzer instrument within 5 min.

*G2938-90025* *G2938-90025* G2938-90025 Rev. D Edition 01/2018

For Research Use Only Not for use in diagnostic procedures

Printed in Germany

Agilent Technologies, Inc. 2002-2018 Agilent Technologies

Hewlett-Packard-Strae 8 76337 Waldbronn, Germany

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