Thermo Fisher Scientific VetMAX S Instruction Manual
SUPPLEMENTAL INSTRUCTIONS
VetMAX™ S. ent
erica spp. Kit
Nucleic acid purification protocols validated (according to PCR methods NF U47-600) and optimized for
use with the kit (Cat. No. SALMSPP, SALMSPP50)
Pub. No. MAN0019475 Rev. A.0
Species Sample matrices Test type
• Wipes and boot swabs
•
Poultry
Bovine
WARNING! Read the Safety Data Sheets (SDSs) and follow the handling instructions. W
clothing, and gloves. Safety Data Sheets (SDSs) are available from thermofisher.com/support.
WARNING! BIOHAZARD
applicable local, state/provincial, and/or national regulations for working with biological samples.
. Read the biological hazard safety information at this product’s page at thermofisher.com. Follow all
Feces, slurry, and manure
• Bedding, bottom cage compartments, and soil
• Organs and embryos
• Organs (placental biopsy, fetal spleen)
•
Placental and cervical swabs
• Liquids (fetal stomach contents, vaginal mucus)
• Feces
• Milk
ear appropriate protective eyewear,
Individual
Individual
P
urpose of this guide ....................................................................................... 2
■
Sample selection .......................................................................................... 2
■
Sample storage ........................................................................................... 2
■
Required materials not supplied .............................................................................. 3
■
Recommended DNA purification protocols ...................................................................... 5
■
Procedural guidelines ...................................................................................... 5
■
Purify nucleic acid using the MagMAX™ CORE Nucleic Acid Purification Kit (automated method) ........................... 6
■
Prepare samples for purification with other kits ................................................................. 10
■
Purify nucleic acid using the MagVet™ Universal Isolation Kit (automated method) ...................................... 12
■
Purify DNA using the QIAamp™ DNA Mini Kit (manual method) ..................................................... 13
■
Purify DNA using the NucleoSpin™ Tissue kit (manual method) ..................................................... 15
■
Purify DNA using the NucleoSpin™ 8 Tissue/NucleoSpin™ 96 Tissue kit (manual method) ................................. 16
■
Good laboratory practices for PCR and RT-PCR ................................................................ 17
■
Appendix A Purification with the KingFisher™ Duo Prime or KingFisher™ mL instrument
Required materials not supplied ............................................................................. 18
■
Purification procedure ..................................................................................... 19
■
Appendix B Documentation and support
Customer and technical support ............................................................................. 19
■
Limited product warranty ................................................................................... 20
■
For Laboratory Use.
Purpose of this guide
This guide describes Salmonella enterica spp. bacterial DNA purification protocols that have been validated (according to PCR methods
NF U47‑600) and optimized for downstream use with the Applied Biosystems™ VetMAX™ S. enterica spp. Kit (Cat. No. SALMSPP,
SALMSPP50).
• Automated nucleic acid purification is performed using one of the following instruments: KingFisher™ Flex, MagMAX™ Express-96,
KingFisher™ mL, or KingFisher™ Duo Prime.
• Manual nucleic acid purification uses silica-based spin columns.
Sample selection
Sample type Type of analysis Quantity required and sampling equipment
Abortion application
Organs Individual 20 mg
Placental and cervical swabs Individual 200 μL of eluate collected from a 20–25 cm cotton swab
Liquids Individual
Poultry application
Wipes Individual 20–60 g
Boot swabs Individual 20–60 g
Dust Individual 20–40 g
• 200 μL of fetal stomach contents
•
200 μL of vaginal mucus
Feces, slurry, and manure Individual 50–100 g
Bottom cage compartments, crate, and
bedding
Embryos Combination 30 embryos maximum
Organs Combination 10 similar organs maximum
Livestock‑monit
Feces Individual 50–100 g
Milk Individual 200 μL minimum
oring application
Individual Five, 30 cm × 30 cm units
Sample storage
Sample type Storage
Organs (placental biopsy, fetal
spleen)
Placental and cervical swabs After collection, maintain the samples at 2°C to 8°C until use (up to 48 hours).
Liquids (fetal stomach contents,
vaginal mucus)
• Wipes
• Boot swabs
• Dust
• Slurry and manure
• Bottom cage compartment,
crate, and bedding
After collection, use the fresh samples immediately or store at 2°C to 8°C until use (up to 8 days).
Aft
er use or after 8 days, store samples below −16°C for up to 1 month, or below −70°C for long‑term storage.
After elution of the swab, use the eluate directly to perform nucleic acid purification. After use or after 48 hours,
store samples below −16°C for up to 1 year, or below −70°C for long‑term storage.
After collection, maintain the samples at 2℃ to 8℃ until use (up to 8 days).
After use or after 8 days, store samples below −16°C for up to 1 year, or below −70°C for long‑term storage.
After collection, use the fresh samples immediately or store at 2°C to 8°C until use (up to 8 days).
Aft
er use or after 8 days, store samples below −16°C for up to 1 month, or below −70°C for long‑term storage.
2 VetMAX
™
S. enterica spp. Kit Supplemental Instructions
Sample type Storage
Embryos and organs After sampling, store as indicated:
• Store samples at 2°C to 8°C if the analysis is to be performed within 24 hours of sampling.
• Store samples below −16°C if the analysis is to be performed more than 24 hours after sampling.
After use or after 24 hours, store samples below −16°C for up to 1 year, or below −70°C for long‑term storage.
Feces After collection, maintain the samples at 2°C to 8°C until use (up to 8 days).
Aft
er use or after 8 days, store samples below −16°C for up to 1 month, or below −70°C for long‑term storage.
Milk After collection, immediately add preservatives, then maintain the samples at 2℃ to 8℃ until use (up to 8 days).
After use or after 8 days, store samples below −16°C for up to 1 year, or below −70°C for long‑term storage.
Required materials not supplied
Unless other
fisherscientific.com or another major laboratory supplier.
Materials required for sample collection, preparation, and nucleic acid purification
Table 1 Materials required for all sample preparation methods
Equipment
Type II Biological Safety Cabinet (BSCII) MLS
wise indicated, all materials are available through thermofisher.com. "MLS" indicates that the material is available from
Item Source
Benchtop microcentrifuge MLS
Laboratory mixer, vortex or equivalent MLS
Adjustable precision micropipettors (range of 1 μL to 1,000 μL) MLS
Consumables
Aerosol-resistant, nuclease-free pipette tips MLS
1.5‑mL and 2.0‑mL DNase/RNase-fr
Reagents
5 – IPC Salmo
Nuclease-free water AM9932
PBS (1X), pH 7.4 MLS
Table 2 Additional materials required for purification fr
Equipment
(Optional) T
• Fisher Scientific™ Bead Mill 24 Homogenizer
• Precellys™ 24 Homogenizer (Bertin)
• FastPrep‑24™ Instrument (MP Biomedical 116004500)
• Mixer Mill 400 (Verder 207450001)
issue homogenizer for bead-beating, one of the following, or equivalent:
ee microtubes MLS
From the VetMAX™ S. enterica spp. Kit
(Cat. No. SALMSPP, SALMSPP50)
om organ samples
Item Source
•
Fisher Scientific 15-340-163
• Bertin EQ03119.200.RD000.0
• Fisher Scientific MP116004500
• Fisher Scientific 08‑418‑241
Precision scale MLS
PYREX™ Solid Glass Beads for Distillation Columns (3 mm), or equivalent 3‑mm glass beads Fisher Scientific™ 11-312-10A
Scalpels and metallic forceps (sterile) MLS
Consumables
Petri dish (sterile) MLS
VetMAX™ S. enterica spp. Kit Supplemental Instructions 3
Table 3 Additional materials required for sample enrichment (for poultry and livestock‑monitoring applications)
Item Source
Equipment
Stomacher™ Model 3500 Jumbo Lab Blender, or equivalent peristaltic laboratory blender Fisher Scientific 14‑285‑32
Stomacher™ 400 homogenizer bag rack, or equivalent Fisher Scientific 14‑285‑3
Stomacher™ 400 homogenizer bag clips, or equivalent Fisher Scientific 13‑874‑850
Consumables
Stomacher™ 400 homogenizer bags, or equivalent Fisher Scientific 14‑285‑24
Reagents
Buered Peptone Water (BPW)
CM0509T
Additional materials required for automated nucleic acid purification
Table 4 Materials required for the MagMAX™ CORE Nucleic Acid Purification Kit
Item Source
Instrument, one of the following:
KingFisher™ Flex Purification System
MagMAX™ Express‑96 Magnetic Particle Processor
KingFisher™ Duo Prime Purification System
KingFisher™ mL Purification System
Equipment
Heat block at 55°C MLS
Reagent reservoir MLS
Consumables
Adhesive PCR Plate Foils, or equivalent AB0626
Consumables for the KingFisher™ Flex and MagMAX™ Express-96 instruments:
• KingFisher™ Deepwell 96 Plate
• KingFisher™ 96 KF microplates
• KingFisher™ 96 tip comb for DW magnets
Contact your local sales oce.
• 95040450
•
97002540
• 97002534
Consumables for the KingFisher™ Duo P
Kits and reagents
MagMAX™ CORE Nucleic Acid Purification Kit A32700 or A32702
PBS, pH 7.4 (10X), RNase‑free AM9624
rime and KingFisher™ mL instruments See Table 9 on page 18.
Table 5 Materials required for the MagVet™ Universal Isolation Kit
Item Source
Instrument, one of the following:
KingFisher™ Fle
KingFisher™ mL Purification System
Equipment
Heat block at 70℃ MLS
Reagent reservoir MLS
4 VetMAX
x Purification System
Contact your local sales oce.MagMAX™ Express‑96 Magnetic Particle Processor
™
S. enterica spp. Kit Supplemental Instructions
Item Source
Kits and reagents
MagVet™ Universal Isolation Kit MV384
Proteinase K (PK)
Ethanol, 80% MLS
• Qiagen 19131
• Macherey Nagel 740396
Additional materials required for manual nucleic acid purification
Item Source
Equipment
Heat block at 70℃ MLS
Kits and reagents
One of the following kits:
QIAamp™ DNA Mini Kit
•
• NucleoSpin™ Tissue kit
• NucleoSpin™ 96 Tissue kit
• NucleoSpin™ 8 Tissue kit
Ethanol, 96–100% MLS
• Qiagen 51304
•
Macherey Nagel 740952
• Macherey Nagel 740741.4
• Macherey Nagel 740740
Recommended DNA purification pr
Purification pr
MagMAX™ CORE Nucleic Acid Purification Kit (page 6) ✓ — —
MagVet™ Universal Isolation Kit (page 12) ✓ ✓ ✓
QIAamp™ DNA Mini Kit (page 13) ✓ ✓ ✓
NucleoSpin™ Tissue kit (page 15) ✓ ✓ ✓
NucleoSpin™ 8 Tissue/NucleoSpin™ 96 Tissue kit (page 16) ✓ ✓ ✓
otocol
otocols
Application
Abortion Poultry Livestock‑monitoring
Procedural guidelines
repare at least one mock‑purified sample for use as a negative extraction control—use PBS (1X), pH 7.4, or nuclease‑free water in place
P
of the test sample, unless otherwise directed. Process the mock-purified sample concurrently with the test samples, using the same
nucleic acid purification protocol.
VetMAX™ S. ent
erica spp. Kit Supplemental Instructions 5
Purify nucleic acid using the MagMAX™ CORE Nucleic Acid Purification Kit (automated method)
This procedure is designed for rapid purification of bacterial DNA from the following sample types for the abortion application.
• Organs (placental biopsy, fetal spleen)
• Placental and cervical swabs
• Liquids (fetal stomach contents, vaginal mucus)
Follow this procedure if you are using these instruments:
• KingFisher™ Flex
• MagMAX™ Express-96
Follow Appendix A, “Purification with the KingFisher™ Duo Prime or KingFisher™ mL instrument” if you are using these instruments:
• KingFisher™ Duo Prime
• KingFisher™ mL
Workflow
MagMAX™ CORE Nucleic Acid P
urification Kit workflow
Set up the processing plates
Prepare the sample
Prepare Lysis/PK Solution
Treat samples with the Lysis/PK Solution
Combine samples with Binding/ Bead/ IPC Mix
Process samples on the instrument
Procedural guidelines
Before use, invert bottles of solutions and buers to ensure thorough mixing.
•
• To prevent cross-contamination:
– Cover the plate or tube strip during the incubation and shaking steps, to prevent spill-over.
– Carefully pipet reagents and samples, to avoid splashing.
• To prevent nuclease contamination:
– Wear laboratory gloves during the procedures. Gloves protect you from the reagents, and they protect the nucleic acid from
nucleases that are present on skin.
– Use nucleic acid-free pipette tips to handle the reagents, and avoid putting used tips into the reagent containers.
– Decontaminate lab benches and pipettes before you begin.
Before first use of the kit
(Optional) Determine the optimal bead mill homogenizer settings
We recommend using the Fisher Scientific™ Bead Mill 24 Homogenizer for maximum nucleic acid yield. If an alternative instrument is
used, follow the manufacturer's guidelines to determine the speed and time settings necessary to achieve sucient cell lysis.
6 V
etMAX™ S. enterica spp. Kit Supplemental Instructions
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