PCRopsisTM Reagent RVD For In Vitro Diagnostic Use
REF #: 783025 & 783100
Store at room temperature
INTENDED USE PCRopsis™ Reagent RVD is intended for extraction-free amplification of RNA from RNA viruses collected using a nasopharyngeal or oropharyngeal swab and transported in compatible transport mediums within 7 days from collection and properly stored.
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01 INTRODUCTION PCRopsis™ Reagent RVD is engineered to simultaneously bind a variety of reverse transcriptase quantitative polymerase chain reaction (RT-qPCR) inhibitors found in RNA viral samples, lyse RNA viruses in transport medium and stabilize RNA in a manner that's compatible with RTqPCR. The product consists of a proprietary mixture of salts, stabilizers, buffers and sodium azide to achieve this task, without employing detergents. Reagent RVD allows for extraction-free amplification of viral RNA without performing RNA isolation, centrifugations or other sample manipulations, which may introduce errors, contaminants and/or skew the representation of RNA fragments.
02 PRODUCT SIZE
CatalogNumber Volume
783025 25mL
783100 100mL
03 STORAGE & STABILITY
PCRopsis™ Reagent RVD is shipped and stored at room temperature. The recommended storage temperature is: 4°C ~ 25°C
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04 TRANSPORT MEDIUM COMPATIBILITY
RECOMMENDED:
• PCRopsis™ Universal Viral Transport (UVT) – preferred
• MedSchenker Smart Transport Medium - preferred
• BD™ Universal Viral Transport System (UVT)
• Quest V-C-M Medium
• Puritan UniTranz-RT Universal Transport Solution
• Remel MicroTest™ M6™ Transport Medium
• TransPRO CVM™ Transport Medium
• Mediums with recipes similar to PCRopsis™ UVT are expected to be compatible
• Copan ESwab™ Amies Medium
NOT RECOMMENDED:
u Phosphate buffered saline (PBS)
u Mediums containing guanidinium thiocyanate
u Hank's Balanced Salt Solution (HBSS) + 2% Fetal Bovine Serum (FBS)
u Water with veal infusion broth + BSA + antibiotics
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05 OVERVIEW OF PROTOCOL
Reagent RVD
Viral test sample in Universal Viral
Transport medium
Mix 50 μL Reagent RVD + 50 μL Sample /
well
Heat Reagent RVD + Sample mixture to 95°C for 10 minutes
Use 5 μL of processed sample into your desired RT-qPCR mixture
NOTE: samples can be heated in a
thermal cycler or heating block
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06 WRITTEN PROTOCOL
1. Thoroughly mix Reagent RVD to ensure homogeneity, but avoid creating bubbles unnecessarily
1. Reagent RVD has a hazy, white color when homogenized and normal settlement occurs if
not regularly mixed
2. Mix 1 volume of sample (50 μL) with 1 volume of Reagent RVD (50 μL) in a thin walled tube (0.2 ~
0.6 mL) or 96-well PCR plate
3. Pipette up & down to ensure complete mixing and then cap tube or apply plate sealer to plate
to prevent evaporation
4. Heat diluted sample for 10 minutes at 95°C and let cool at room temperature for ~10 seconds
before continuing
1. Alternatively, you can heat the diluted sample for 15 minutes at 80°C
2. Make sure the heating device has reached the desired temperature before applying
sample
3. Sample heating can be performed using a controlled heating block or thermal cycler
5. Mix heated sample and use 1 - 5 μL of lysed / stabilized sample in your desired RT-qPCR
procedure
1. 25% of lysed sample can be added to your RT-qPCR mixture (i.e., 5 μL sample into a total
volume of 20 μL)
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07 STEP-BY-STEP PROTOCOL WITH FIGURES
Step 1
Gently invert Reagent
RVD to ensure
homogeneity
Step 2
Add Reagent RVD to
reservoir
Step 3
Add 50 μL of Reagent
RVD to wells in a 96-
well PCR plate PCRopsisTM Reagent RVD 6 of 10
Step 4 Step 5
Seal 96-well plate with
a plate sealer to
prevent evaporation
Step 6
Mix 50 μL of viral sample
to wells containing 50 μL
of Reagent RVD
07 STEP-BY-STEP PROTOCOL WITH FIGURES
Heat Reagent RVD +
sample mixture to 95°C
for 10 minutes
Pre-heated thermal cycler or heating block before
applying plate
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Step 7 Step 8
Mix 5 μL of heated Reagent
RVD + sample mixture with
your RT-qPCR mix
Step 9
Add ~15 μL RT-qPCR mix
from your desired vendor
to a new plate
07 STEP-BY-STEP PROTOCOL WITH FIGURES
Detect amplification of
target genes using your
desired qPCR equipment PCRopsisTM Reagent RVD 8 of 10
08 TROUBLESHOOTING & SUGGESTIONS
1. Reagent RVD is optimized for the amplification of gene targets from RNA viruses in compatible
transport mediums and may not be applicable for other applications or mediums.
2. For best results, use recently collected viral samples in compatible transport mediums that
have been stored at ~4°C since collection.
3. Samples are diluted 50% with Reagent RVD. As such, you should expect to observe slightly
higher Ct’s compared to RNA extracted samples.
4. Ct cut-offs for assays should be increased, often times to 40 cycles.
5. Take care in maintaining the sterility of your Reagent RVD stock after use.
6. It’s recommended to immediately use the heated Reagent RVD + sample mixture for
downstream applications.
09 CONTACT
Contact our research team if assistance with Reagent RVD is necessary ([email protected]). We
will try our best to assist with non-intended applications of this product or direct you to alternative
products. Any business related questions should be directed to: [email protected].
PCRopsisTM Reagent RVD 9 of 10
7600 NW 69th Ave, Medley, FL 33166
www.PCRopsis.com
Toll-free 1-888-407-5070
PCRopsisTM is a trademark of Entopsis
v.20200827
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