InstantLabs Listeria species Food Safety Kit
Contents
1. e MAC Assay Cartridges MAC Additional Components e Hunter device User Manual CD e 2 spare fuses e AC line cord Hunter instrument specifications e 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 lbs 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 e 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 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 reserved2. 6 0 f ININ IN INSTANTLABS LABORATORY USE ONLY InstantLabs Listeria species Food Safety Kit P N 9034 0600 0015 PRODUCT DESCRIPTION The InstantLabs Medical Diagnostics Corporation InstantLabs Listeria species Food Safety 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 Food Safety Kit at 2 8 C The reagents are stable through the expiration date printed on the kit MATERIALS REQUIRED AND PROVIDED e Multiple Assay Cartridges MAC e Reagent Packs each containing the following o 5 Colored Tubes With Lyophilized PCR Reagents 4 Sample Extraction Tubes 1 Tube of PCR Positive Control 1 Tube of Nuclease Free Water O O O ADDITIONAL MATERIALS REQUIRED BUT NOT PROVIDED 1 Hunter Accelerated PCR Instrument 2 Nitrile or Latex Gloves 3 Buffered Listeria Enrichment Broth InstantLabs or Other Supplier 10 100 uL Pipette with RNase DNase Free Filter Tips Heat Block Set at 95 C Incubator Set at 35 C Micro Centrifuge Capable of at Least 6 000 x g Vortex w 1 5 mL Tube Adaptor 2 a For Food Samples Only 9 Whirl Pak Filter Bag for Homogenizer Blenders or Other Similar Filter Bag 10 Sterile Transfer Pipet of Suitable Capacity Or S
3. A Extraction from Enriched Samples 1 Clean the work area and laboratory pipets with a 10 bleach solution and wipe until dry Firmly tap Sample Extraction tubes on the laboratory bench to collect contents at bottom of tube Label each tube or complete included tracking card with identifying information for the sample to be tested Remove Whirl Pak bag s or tube s from incubator and gently agitate the contents Allow large food debris to settle For Food Samples Carefully open the bag and remove 25 uL of liquid from the top 1 3 cm of the filtered compartment of the Whirl Pak bag without disturbing the settled food debris and place into the correctly labeled Sample Extraction tube prepared in step 2 above Note that if a layer of fat is present move pipet tip to below the fat layer before removing aliquot For Environmental Samples Carefully open the bag or tube and remove 25 uL of liquid and place into the correctly labeled Sample Extraction tube prepared in step 2 above Repeat step 4 for each sample to be tested Gently invert Sample Extraction tube s 3 5 times to ensure sample is mixed with the lysis reagents Vortex Sample Extraction tubes at maximum speed for 10 minutes Use a 1 5 mL tube adaptor on the vortex to securely hold tube 8 Heat Sample Extraction tubes to 95 C 1 C for 10 minutes in a dry heat block to complete the lysis Allow tube s cool at room temperature for 1 2 minutes 9 Gen
4. ATION The InstantLabs Listeria species Food Verification Kit targets a specific region of bacterial genomic DNA which can be present in high copy number in contaminated foods 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 and laboratory balances by following the manufactures instructions Always use a 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 SAMPLES 25 g Sample Collec
5. e until dry Using clean gloves remove Lyophilized PCR Reagents Pack from the kit Make sure the lyophilized reagents are collected in the bottom of each tube by tapping tubes on benchtop Using a sterile pipet transfer 25 uL of Nuclease Free Water to the blue colored Negative Control tube Tap tube gently on benchtop to ensure that lyophilized reagents are reconstituted Transfer 25 uL of the extracted DNA sample to be tested to the green colored tube using a sterile pipet Tap tube gently on benchtop to ensure that lyophilized reagents are reconstituted Additional extracted samples up to a total of 4 can be similarly loaded into the Orange Purple and Yellow tubes 10 11 12 13 if desired Tap the tubes gently on benchtop to ensure that lyophilized reagents are reconstituted Using a sterile pipet transfer 25 uL of Nuclease Free Water to the red tube labeled PCR Positive Control Tap tube gently on benchtop to ensure that lyophilized reagents are reconstituted Flick each colored tube with your finger to ensure reagents are dissolved and properly mixed and allow to incubate for 2 3 minutes before moving on to the next step Do NOT vortex this may cause excess foaming Briefly spin down the 6 colored tubes for approximately 15 seconds to collect contents at bottom of tube Remove one MAC from the kit Prepare Hunter instrument by entering test and sample information as outlined in the Hunter User Manua
6. imilar Sterile Liquid Measuring Device Liquid Food Samples Only 11 Sterile Spatula Solid Food Samples Only 12 Laboratory Balance with a Sensitivity of at Least 1 g Solid Food Samples Only 13 Stomacher Instrument Or Similar Homogenizer Blender Optional for most food types but recommended for meat and other samples that cannot be effectively mixed by hand For Environmental Surface Samples Only 14 Sterile Swabs Pre Moistened with Neutralizing Buffer Sealed Concrete Surfaces Only 15 Sterile Sponges Pre Moistened with Neutralizing Buffer Ceramic and Stainless Steel Surfaces Only 16 Sterile 15 ml capped tubes Swabs Only 17 Whirl Pak Bags Non Filtered Sponges Only WARNINGS AND PRECAUTIONS 1 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 excess reagents from one reagent pack to another dispose of all remaining reagents once all kit MACs have been used Material Safety Data Sheets MSDS are available upon request Treat enriched samples as if they contain pathogenic bacteria Enriched samples and MACs should be disposed of as bio hazardous waste according to local regulations AVOIDING CROSS CONTAMIN
7. ion and Enrichment SURFACE TYPE AREA MEDIA USED Sealed Concrete Swab 1 x 1 10 ml Stainless Steel Sponge 4 x 4 100 ml 1 Prepare the designated amount of Buffered Listeria Enrichment Broth BLEB for each sample to be tested by following the manufacturer s instructions see above table for required volumes Bring the media to approximately room temperature immediately before use For swab samples aliquot 10 ml of BLEB to the appropriate number of sterile 15 ml tubes 2 Depending on the surface to be tested wipe the specified area with the pre moistened swab or sponge see above table Wipe the entire area side to side then repeat the motion at a 90 angle making sure to wipe the entirety of the area specified 3 Prepare sample for enrichment a For swabs break off the top of the swab into the 15 mL tube containing 10 ml of BLEB and cap Vortex for 15 30 seconds to dislodge bacterial cells Loosen cap slightly b For sponges place the entire sterile portion of the sponge in the Whirl Pak Bag along with 100 ml of sterile BLEB Close and vigorously hand massage for 2 minutes Jf the entirety of the sponge sponge handle is not sterilized break off or sequester the non sterile portion of the sponge sponge handle Repeat steps 2 and 3 above for each additional sample to be tested Incubate the Whirl Pak bag s without shaking at 35 C 1 C for 22 28 hours DN
8. l Pipette 20u1 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 Ensure 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 It is recommended that the Negative Control is loaded first followed by the samples and finally the Positive Control 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 v lt UUE 14 Place the MAC into Hunter and firmly push latch down to lock MAC in place 15 Close the Hunter s door to start the test which will begin in 15 seconds 16 The results will be displayed automatically as Positive Negative or Indeterminate when the test is completed TROUBLESHOOTING Problem Suggestion e Positive Control was expired not reconstituted properly or was not loaded into the MAC correctly Repeat test with fresh reagents e Kit was not stored correctly No amplification of Positive Control The Hunter calls the positive control Negative or Indeterminate Repeat test with fresh reagents High background in the e Potential contamination Repeat Negative Control test with fresh reagents Hunter calls all samples indeterminate 7 HUNTER SYSTEM SPECIFICATIONS System Components e Hunter device
9. tion Homogenization and Enrichment 1 Prepare 225 mL of Buffered Listeria Enrichment Broth BLEB for each sample to be tested by following the manufacturer s instructions Ensure that the temperature of the broth is at approximately room temperature immediately before use Clean the work area and weighing scale with a 10 bleach solution and wipe until dry Using clean disposable gloves carefully open and place a new Whirl Pak bag onto the laboratory scale Aseptically transfer approximately 25 g of a representative food sample to the bag using a sterile spatula or scoop For liquid samples transfer 25 mL using a sterile pipet It is important to try to work quickly to minimize the chances of contaminating the sample Repeat steps 3 and 4 above for each additional sample to be tested Pour 225 mL of room temperature BLEB into each bag and loosely seal bag with built in wire closures f necessary sample size may deviate from 25 g depending on availability of the sample as long as the amount of BLEB is adjusted proportionally Gently hand massage Whirl Pak bag s for 1 2 minutes to mix contents Note that samples containing meats or other difficult to mix materials should instead be homogenized using a Stomacher machine for 30 60 seconds at normal speed 8 Incubate the Whirl Pak bag s without shaking at 35 C 1 C for 24 28 hours FOR TESTING ENVIRONMENTAL SURFACE SAMPLES Sample Collection Homogenizat
10. tly invert Sample Extraction tube s 3 5 times and then centrifuge tube s in a Micro Centrifuge for 2 minutes at maximum speed to pellet the lysing beads and any food debris The supernatant contains the extracted DNA and is ready to be tested immediately with the InstantLabs Food Safety Kit see next section on PCR Setup Note that when removing the supernatant be careful to avoid disturbing settled material For food samples with high fat or oil content an oil fat layer can form over the extracted DNA sample in the Sample Extraction tube Avoid the top layer and collect the sample from below for PCR Supernatant Eee lt Contains DONA oc y Avoid Beads and Food Debris Graphic of Sample Extraction Tube Following Centrifugation Avoid Fat Layer If Present If sample testing will be delayed beyond 30 minutes transfer 100 uL of supernatant to a new nuclease free microcentrifuge tube and freeze at 20 C The samples may be frozen for up to 3 months prior to testing Thaw frozen samples on ice and centrifuge at maximum speed for 3 minutes before use 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 wip
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