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Agilent Protein 80 Kit Quick Start Guide

1. Agilent Protein 80 Kit e o o e oe e ec Agilent Protein 80 Kit 909 Quick Start Guide 6 Protein 80 Kit reorder number 5067 1515 Protein Chips Protein 80 Reagents reagent reorder number 5067 1516 amp Supplies 25 Protein Chips red Protein 80 Gel Matrix 4 vials in box labelled Part Store at 4 C 1 Electrode Cleaner blue Protein 80 Dye Concentrate in box labelled Part Store at 4 C Oj white Protein 80 Sample Buffer 4 vials in box labelled Part II Store at 20 C Syringe Kit yellow Protein 80 Ladder in box labelled Part II Store at 20 C 1 Syringe 4 Spin Filters This product is provided under an agreement between Molecular Probes Inc a wholly owned subsidiary of Invitrogen Corporation and Agilent Technologies The manufacture use sale or import of this product may be subject to one or more of U S patents pending applications and corresponding international equivalents owned by Molecular Probes Inc The purchaser has the non transferable right to use the product to detect protein and or nucleic acids in microfluidics analysis systems for one or more of the subfields of research development quality control forensics environmental analysis biodefense food safety testing veterinary diagnostics or human diagnostics according to use indicated on the product label or accompanying product literature For information on obtaining a license contact Molecular Probes
2. Inc Business Development 29851 Willow Creek Road Eugene OR 97402 9132 Tel 541 465 8300 Fax 541 335 0354 Assay Principles Agilent Protein kits contain chips and reagents designed for sizing and analysis of proteins Each chip contains an interconnected set of gel filled microchannels that sieves proteins by size as they are driven through it by means of electrophoresis Agilent Protein kits are designed for use with the Agilent 2100 bioanalyzer only Assay Kits The Agilent Protein 80 kit is designed for the sizing and analysis of proteins from 5 80 kDa and can be used to analyze cell lysates column fractions or purified proteins The complete Protein 80 Kit Guide can be found in the online help of the 2100 expert software Other protein kits from Agilent Protein 230 kit reorder number 5067 1517 Storage Conditions e Keep all reagents in box labelled Part I refrigerated at 4 C when not in use to avoid poor results caused by reagent decomposition e Store Protein 80 Sample Buffer and Ladder box Part II at 20 C upon arrival To avoid freeze thaw cycles make aliquots depending on your daily use e g 6 ul for ladder The aliquot in use should be stored at 4 C e Protect all reagents from light Remove light covers only when pipetting The reagents contain dye that decomposes when exposed to light fee Agilent Technologies Agilent Protein 80 Kit Quick Start Guide Equipment Supplied with the Agilent 2100 B
3. atrix vial red to a spin filter Make sure the complete volume of 650 pl has been transferred 2 Centrifuge at 2500 g 20 for 15 min 77 destaining solution 650 pl gel 3 Label with the date and DS Destaining Solution Use within kit life time Preparing the Denaturing Solution 1 For reducing conditions add 3 5 Vol of 1 M Dithiothreitol DTT or mercaptoethanol BME to an aliquot of sample buffer e g 1 0 ul DTT or BME to an aliquot of 28 6 ul Sample Buffer Alternatively for non reducing conditions add 3 5 Vol of water to your aliquoted sample buffer vial 2 Vortex for 5s 7 7 denaturing solution e g 1 0 pl DTT 28 6 pl sample buffer Preparing the Samples and the Ladder 1 Combine 4 ul protein sample and 2 pl denaturing solution in a 0 5 ml vial 2 Place sample vial and a vial containing a 6 ul aliquot of Protein 80 Ladder yellow at 95 C for 5 min Cool down afterwards 3 Spin tubes for 15 s 4 Add 84 ul deionized water to samples and ladder and vortex 77 2 pl denaturing solution 4 ul sample 84 ul deionized water J Loading the Gel Dye Mix and the Destaining Solution 1 Adjust the base plate of the chip priming station to position A and the syringe clip to its middle position Take a new protein chip out of the sealed bag and put it on the chip priming station Pipette 12 ul of gel dye mix in the well marked Put plunger at 1 ml and close chip priming station Press plunger
4. e loaded chips within 5 minutes Reagents might evaporate leading to poor results Do not touch the Agilent 2100 bioanalyzer during analysis and never place it on a vibrating surface Use 0 5 ml vials to denature samples Using larger vials may lead to poor results caused by evaporation Agilent Protein 80 Assay Protocol Edition April 2007 Handling DMSO Some solutions may contain DMSO dye 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 dye solutions with particular caution as DMSO is known to facilitate the entry of organic molecules into tissues Preparing the Gel Dye Mix 1 l l Pa 77 gel dye mix Transfer the content 650 pl of an Agilent Protein 80 Gel matrix vial red to 25 pl dye a spin filter Make sure the complete volume of 650 ul has been transfered 650 pl gel Centrifuge at 2500 g 20 for 15 min To the filtered and centrifuged Gel add 25 ul of the well vortexed Dye concentrate blue Mix thoroughly for 10 20 s Vortexer until an uniform color is obtained Label with the date and G D Gel Dye Use within 4 weeks Agilent Protein 80 Kit Quick Start Guide Agilent Protein 80 Assay Protocol Edition April 2007 Destaining Solution 1 Transfer the content 650 ul of another Protein 80 Gel m
5. ide 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 A Retighten the screw 3 Agilent Protein 80 Kit Quick Start Guide Adjust the syringe clip a Release the lever of the clip and lift it up or down to adjust it to the middle 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 Upon arrival make aliquots of sample buffer and ladder with the typical amount required for daily use and store them at 20 C Keep the vial in use at 4 C to avoid freeze thaw cycles Allow all reagents and samples to equilibrate to room temperature for 30 minutes before use Protect all reagents from light Remove light covers only when pipetting The dye contained in the reagents 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 Us
6. ioanalyzer e Chip priming station reorder number 5065 4401 Additional Material Required Not Supplied e Pipettes 10 ul 20 ul 100 ul and 1000 ul with e 1M Dithiothreitol DTT solution or compatible tips 2 Mercaptoethanol BME e 0 5 ml microcentrifuge vials e Microcentrifuge e Deionized water e Heating block or water bath for 0 5 ml vials Physical Specifications Analytical Specifications Type Specification Type Agilent Protein 80 Assay Analysis runtime 30 minutes Sizing range 5 80 kDa Number of samples 10 samples chip Typical sizing resolution 10 Sample volume 4ul Typical sizing accuracy 10 CV CAII BLG Kit stability 4 months Storage Temperature Sizing reproducibility 3 CV CAII BLG see individual box Sensitivity 6 ng ul CAII 15 ng pl BSA in PBS 10 ng ul Signal Noise gt 3 CAII in 0 5 M NaCl 30ng pl BSA in 0 5 M NaCl CAII Carbonic Anhydrase Quantitative range 60 2000 ng pul CAII in PBS BSA Bovine Serum Albumin Qualitative range 6 4000 ng pl CAII and BLG BLG beta Lactoglobulin Quantitation reproducibility 20 CV CAII BLG Compatible buffers see List of Compatible Buffers and Buffer Compounds in your Protein 80 Kit Guide Setting up the Chip Priming Station 1 Replace the syringe a Unscrew the old syringe from the lid of the chip priming station Release the old syringe from the clip Discard the old syringe b c Remove the plastic cap of the new syringe and insert it into the clip d Sl
7. until held by clip wait 60 s then release clip Wait for 5 s Slowly pull back plunger to 1ml position Pipette 12 ul of gel dye mix in all wells labeled with G con amp C1 F amp F GW NO Loading the Ladder and the Samples 1 Pipette 6 pl of sample in all 10 sample wells Note all 10 sample wells must be filled either with ladder or sample 2 Pipette 6 pl of the prepared ladder in the well marked 3 Place the chip in the Agilent 2100 bioanalyzer and start the assay immediately Pressurize oo0oo0oo0 oo0oo0oo0o okok ok 12pl destain Pipette 12 ul of destaining solution in well DS 6yl sample Technical Support In the U S Canada 1 800 227 9770 toll free Isca ibs support agilent com In Europe call your local Customer Care Center bio_solutions agilent com In Japan 0120 477 111 yan_ccr agilent com In Asia Pacific call your local Customer Care Center Bioanalyzer_ap agilent com Further Information Visit Agilent Technologies unique Lab on a Chip web site It is offering useful information support and current developments about the products and the technology http www agilent com chem labonachip AA AA A TUIT Part Number 62938 90063 Agilent Technologies Inc 2000 2001 2007 MUNTEN ON N VI TTT Edition 04 2007 Agilent Technologies Deutschland GmbH G2938 90063 Printed in Germany Hewlett Packard Stralhe 8 76337 Waldbronn Germany

Agilent Protein 80 Kit Quick Start Guide

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