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InstantLabs Porcine Species Food Verification Kit
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1. is loaded in QuickGene instrument Be careful to avoid transferring any pelleted food debris Process samples according to the standard QuickGene protocol including 2 washes with Buffer WDT and elution in 200 uL of Buffer CDT Note the following o Ensure that Ethanol 299 has been added to Buffer WDT before use as described in the QuickGene manual o Make sure that no WDT remains in the cartridge before beginning the elution step o Incubate columns at room temperature for 90 seconds after adding Buffer CDT before applying pressure to elute DNA The eluted DNA is now ready for testing immediately with the InstantLabs Food Verification Kit see below or can be stored at 20 C for up to 3 months if desired for later analysis PCR Setup Overview Each Reagent Pack and Multiple Assay Cartridge MAC can be used to run up to 4 samples plus the required positive and negative controls The tubes in the reagent pack are color coded to match the wells on the MAC and contain convenient ready to use lyophilized reagents Procedure 1 Clean workspace and pipets with a 10 bleach solution and wipe until dry Using clean gloves remove one Reagent Pack from the kit Transfer 25 uL of the extracted DNA sample to be tested to the green colored tube using a sterile pipet while being careful to not touch the lyophilized reagents with the pipet tip Shake tube gently to ensure that lyophilized reagents are reconstituted Add
2. 6 0 f ININ IN INSTANTLABS LABORATORY USE ONLY InstantLabs Porcine Species Food Verification Kit P N 9034 0600 0012 PRODUCT DESCRIPTION The InstantLabs Medical Diagnostics Corporation InstantLabs Porcine Species Food Verification Kit contains the reagents necessary for performing qualitative real time PCR on the Hunter Accelerated PCR system see Hunter System Specifications below The kit is intended to be used in an analytical laboratory under standard laboratory conditions KIT STORAGE AND STABILITY Store the InstantLabs Porcine Species Food Verification Kit at 2 8 C The reagents are stable through the expiration date printed on kit MATERIALS REQUIRED AND PROVIDED e Multiple Assay Cartridges MAC e Reagent Packs each containing the following o 6 Colored Tubes With Lyophilized PCR Reagents o 1 Tube Positive Control DNA o 1 Tube Nuclease Free Water ADDITIONAL MATERIALS REQUIRED BUT NOT PROVIDED SO RST OY Beh Hunter Accelerated PCR Instrument InstantLabs Extraction Buffer P1 InstantLabs 10 Bleach Solution Nitrile or Latex Gloves FujiFilm QuickGene Mini 80 Instrument InstantLabs FujiFilm QuickGene DNA Tissue S Kit InstantLabs Vortex Mixer 20 200 uL Pipette with RNase DNase Free Filter Tips 100 1000 uL Pipette with RNase DNase Free Filter Tips Sterile 50 mL Conical Tubes RNase DNase Free Heat Block Set at 55 C With Insert for 50 mL Conical Tubes Micro Centrifu
3. ge Capable of at Least 6 000 x g Laboratory Balance With Minimum 0 1 g Sensitivity Ethanol 99 1 5 mL Microcentrifuge Tubes RNase DNase Free Sharpie Marker or Similar to Label Tubes WARNINGS AND PRECAUTIONS il Wear eye protection gloves and laboratory coats when handling samples and kit reagents Avoid contamination of reagents by only using sterile pipette tips Do not use kits beyond the printed expiration date Do not use kits that have been stored above 8 C A negative and a positive control must be included in each test Do not reuse un used or partial reagents from one kit to another dispose of all remaining reagents once all kit MACs have been used Material Safety Data Sheets MSDS are available upon request Samples and MACs should be disposed of as bio hazardous waste according to local regulations AVOIDING CROSS CONTAMINATION The InstantLabs Porcine Species Food Verification Kit targets a region of the mitochondrial DNA that can be present in high copy number in foods and other materials that are produced with porcine derived materials To minimize the potential for cross contamination of samples and false positive results please review these recommendations A Always wipe the bench area with a freshly prepared 10 bleach solution and wipe until dry before and after running the assay Regularly clean and decontaminate pipets tube racks centrifuge laboratory balance and the Q
4. ght to each tube For example if the weight of the sample is I g then add 10 mL of Extraction Buffer P1 Add 20 uL of Buffer EDT to each tube and vortex for 30 seconds to mix Note Buffer EDT is a component of the QuickGene DNA Tissue S kit Incubate sample tube s at 55 C for 60 minutes with occasional mixing on the vortex to dissolve Note that sample may not completely dissolve Let the sample tubes sit undisturbed at room temperature for 3 5 minutes This brief incubation allows the sample to cool and for any fats or oils present in the sample to collect at the top of the solution above the lower DNA containing aqueous phase The dissolved samples are now ready for the DNA Extraction step DNA Extraction 1 6 Using clean gloves carefully load the QuickGene instrument with appropriate plastic ware including fresh columns collection tubes and elution tubes refer to QuickGene user manual for instructions Combine 180 uL of Buffer LDT and 240 uL of Ethanol 99 into a new labeled 1 5 mL tube for each sample Invert tube s 3 5 times to mix Transfer 250 uL of each dissolved sample from step 8 above to the appropriate 1 5 mL tube prepared in the previous step and invert 3 5 times to mix Be careful to avoid the top fat layer if present when transferring the sample Centrifuge 1 5 mL tubes for 3 minutes at maximum speed and then transfer the supernatant using a pipet to the appropriate column that
5. itional lysed samples can be similarly loaded into the Orange Purple and Yellow tubes if desired Shake the tubes to ensure that lyophilized reagents are reconstituted Using a sterile pipet transfer 25 uL of Nuclease Free Water to the blue colored Negative Control tube Shake tube to ensure that lyophilized reagents are reconstituted Using a sterile pipet transfer 50 uL of Nuclease Free Water to the tube labeled Positive Control and invert tube 3 5 times to mix contents Using a new pipet tip transfer 25 uL of this solution to the red colored Positive Control tube Shake tube to ensure that lyophilized reagents are reconstituted 10 11 12 13 14 Briefly spin down the 6 colored tubes in a Micro Centrifuge for 5 seconds to collect contents at bottom of tube Remove one MAC from the kit Prepare Hunter by entering test and sample information as outlined in the Hunter User Manual Pipette 20 ul of each Sample Control PCR mix solution to the correct port on the MAC by matching the color of the tube to the colors printed on the MAC Insure that the solution flows into the sample chamber and then close the cap on the port To ensure a proper seal press the cap down firmly When all samples have been added to the MAC tap the MAC gently on the countertop to ensure there are no visible bubbles present in the sample chamber ig a NO NAIA 2 WOW Place the MAC into Hunter and firmly push latch down to lock MAC i
6. n place Close the Hunter s door to start the test which will begin in 15 seconds The results will be displayed automatically as Positive Negative or Indeterminate when the test is completed TROUBLESHOOTING Problem Suggestion No amplification of Positive Control The Hunter calls the positive control Negative or Indeterminate Positive Control was expired not reconstituted properly stored incorrectly or was not loaded into the MAC properly Repeat test with fresh reagents High background in the Negative Control Hunter calls all samples indeterminate 7 Potential contamination Repeat test with fresh reagents HUNTER SYSTEM SPECIFICATIONS System Components e Hunter device e MAC Assay Cartridges MAC Additional Components e Hunter device User Manual CD e 2 spare fuses e AC line cord Hunter instrument specifications Dimensions 17 2 cm x 41 2 cm x 22 cm 6 77 in H x 16 22 in W x 8 66 in D e Weight 6 8 Kg 15 Ibs e Power usage 100 to 240 VAC at 3 amps e Fuses 2 3 AMP slow blow e Peak block heat rate 4 5 C sec e Peak block cool rate 4 5 C sec e Warm up time 10 minutes e Decibel level less than 40 db at 1 meter Environment Temperature 10 35 C 50 95 F e Relative humidity up to 90 noncondensing e Locate away from heaters cooling ducts and keep out of direct sunlight e Protect from all fluids Throughput e Six 6
7. samples every 2 25 hours run time e Includes pos neg control e 18 samples per 8 hour shift e Supported volumes 20ul Area Requirements e 8 recommended free space around the Hunter system for sample preparation airflow etc TECHNICAL ASSISTANCE Worldwide Support Phone 855 800 7086 Email support instantlabs com Web www instantlabs com 2012 by InstantLabs Medical Diagnostics Corporation Inc All rights reserved DOCUMENT VERSION 10016
8. uickGene instrument by following the manufactures instructions Always use a brand new filtered pipet tip when pipetting reagents or samples Wear clean disposable gloves at all times and change gloves regularly especially between the Sample Preparation DNA Extraction and PCR Setup steps Physically separate the steps of Sample Collection DNA Extraction and PCR Setup into different areas of the laboratory if possible Use a different set of pipets and racks for the Sample Preparation DNA Extraction and PCR Setup if available Use care when opening tubes in order to avoid splashing or spilling of reagents or samples Clean up spills immediately and wash contaminated area with 10 bleach solution and wipe until dry After PCR amplification is complete dispose of the used MAC cartridge appropriately Do not open caps on used MAC cartridge or attempt to reuse INSTRUCTIONS FOR USE FOR TESTING OF FOOD AND NUTRACEUTICALS Sample Preparation 1 Clean the work area and laboratory balance with a 10 bleach solution and wipe until dry Tare a new 50 mL conical tube on the laboratory balance Aseptically transfer approximately 1 g of the sample material to be tested to the empty tube and record the weight Label the tube with identifying information about the sample Repeat steps 2 and 3 for each additional sample to be tested Add a volume of fresh Extraction Buffer P1 that is approximately equal to 10x the sample wei
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