AmpliRun®

AmpliRun®


Vircell PCR Controls have been designed to be used as positive controls in nucleic acid amplification reactions, in both conventional and Real-Time PCR. Moreover, our PCR controls can be used to control the extraction process by adding 1 µl of the PCR control to a negative sample.

Vircell PCR controls contain the complete, purified DNA or RNA of the specified microorganism. Therefore any specific microorganism oligo pair can be used for the amplification of the infectious agent. Only MERS CORONAVIRUS DNA CONTROL (MBC120) does not include the full microorganism DNA. These RNA virus controls are plasmid with a specific gene fragment of the microorganism included and, in the name of the product, its nature of DNA plasmid is specified.

Our PCR controls are supplied in lyophilized presentation. Controls must be reconstituted with sterile bidistilled water. In the case of DNA controls the required volume is 100 µl, while for RNA control only 50 µl are required.

For Vircell PCR controls, DNA can last a maximum of 30 months after its manufacture date while RNA controls can last a maximum of 24 months after its manufacture date.  Always following storage recommendations described in the instructions for use of the product.

Yes, Vircell PCR controls can be run on any platform. The performance of our PCR controls has been proven to remain the same regardless of the platform on which the technique is performed.

No extraction is required prior to use of our PCR controls as they are the complete, purified DNA or RNA of the agent to be analysed. However, as mentioned in the first FAQ, our PCR controls can also be used as extraction controls by adding 1 µl to a negative sample.

Vircell PCR controls are supplied in lyophilized presentation. In this condition they can be stored refrigerated (2-8ºC). Once reconstituted, the DNA controls should be stored at temperatures between -5 and -40ºC, while RNA controls should be stored at temperatures between -70 and -90ºC. Vircell recommends to aliquot the material after reconstitution in order to avoid possible freeze-thawing issues associated with reusing the same vial of material for several tests. The response of the controls after repeated freeze/ thaw cycles has been tested and both DNA and RNA maintain their quality and performance. However, we recommend users avoid unnecessary freezing and thawing in order to prevent degradation or contamination of the controls.

This is not a problem. After reconstituting the product, following the instructions for use, it is possible to make a dilution of the control in order to obtain the concentration that best suits your requirements. However you must be careful, specially with quantified controls, and avoid storing highly diluted aliquots (<1000 copies/µl). Bear in mind that controls at low concentration are less stable to prolonged storage and freeze/ thaw cycles, and the product features can be altered.
Vircell has already quantified most of the controls, to make them specifically suitable for quantitative PCR. Updated information on this regard is available in the product list of our website. Please do not hesitate to contact us for further information regarding our quantified controls.

NOVEL INFLUENZA A H1N1 (plasmid) DNA CONTROL consists of a mixture of two plasmids containing the complete genes of the hemagglutinin and neuraminidase from nH1N1. Therefore, it does not include the complete genetic material of the virus. This control can only be used in PCR, targeting complete or partial sequences of the genes corresponding to these proteins (hemagglutinin or neuraminidase). Although this fact can be seen as a limitation, it enables us to offer a purer and more stable product, with greater lot to lot consistency and it is more easily quantifiable. At present, labs are using a generic retrotranscription step in their RT-PCR assays for the new flu. This step uses random hexamers and is able to reversotranscribe any RNA present in the sample. So the specificity of the reaction is given through the PCR step in which specific primers for the novel H1N1 are used. Our new control is ideally suited to control this PCR step, and it has been developed with this objective in mind; being a DNA plasmid it is not necessary to perform the retrotranscription.

The quantification of our controls has been performed by real-time PCR. The amount of copies has been obtained using a specific microorganism gene as target.

In the quantification by real-time PCR a standard curve is used as a reference by comparing the amplification of a target gene from the microorganism with a specific plasmid standard for each of the references quantified.

In the case of DNA controls the concentration obtained after resuspending the product is 12,500 copies/µl. However, in RNA controls the process of quantification by real time-PCR depends on the retrotranscription step, which can vary depending mainly on the efficiency of the Trasnscriptasa (RT) used. For this reason, in the case of RNA controls we declare a concentration of ≥ 12,500 copies/µl 

As detailed in the instructions for use of the product, it is advisable to store aliquots of the controls to avoid unnecessary freezing/thawing cycles. In the case of requiring a lower concentration of the control, it is possible to make dilutions and stored them for a short period of time. However, our recommendation is never to store dilutions with a concentration below 1000 copies/µl, as after prolonged storage or successive freezing/thawing cycles, a reduction in the number of copies has been observed.

In the case of requiring a lower concentration of the control, it is possible to make dilutions and store them for a short period of time.

We do not recommend freezing and thawing the PCR controls several times as the performance of the control can be affected, especially when the genetic material is not very concentrated (<1000 copies/µl). 

The influence of repeated freezing and thawing cycles in the quality of the product will depend on the kind of PCR control since RNA decomposes more easily than DNA. In any case, it is difficult to set a maximum number of freezing/thawing that guarantees the quality of the process. Thus, we suggest reconstituting the PCR control upon reception and make aliquots before the freezing.

Our PCR controls are not infectious as they only contain the purified genetic material obtained from the previously inactivated microorganism. Vircell provides a document certifying this feature upon customer request.

Vircell recommends using at least 1 µl when first using a PCR control. In quantified controls 1 µl volume contains approximately 12,500 copies of the microorganism genome. Hence, up to 100 runs can be performed with each vial. However, the end-user could prepare dilutions from the control to determine number of copies needed to obtain a clear positive control for the PCR system used. Nevertheless, we do not recommend using the PCR control in the detection limit, which can range from 4 to 10 copies per control.

Vircell PCR controls offer important benefits compared to main competitors: • Wide range of products- more than 120 infectious agents available • Positive controls suitable for conventional and Real Time PCR- any platform can be used • Versatile product line- extraction control and amplification control • Quantified controls available- Vircell is in the process of quantifying most references • Complete purified DNA or RNA- any sequence can be amplified • Obtained from whole microorganisms • Designed to avoid variability factors in PCR testing • Lyophilized presentation avoids extra transport costs and improves stability • No need of dry ice for transport • Competitive price for diagnostic use

Vircell R&D team is currently working on the development of product line extension. In the near future we will offer new controls to validate the nucleic acid extraction and amplification processes. These controls will consist of whole organisms that undergo the same process of extraction and amplification than the clinical sample. For further information, please do not hesitate to contact us.
All our PCR controls, with the exception of the references for Chlamydia, Cytomegalovirus and Toxoplasma are CE marked. The products that are not yet CE marked are under a more complex process and obtaining the CE mark will take more time. Please do not hesitate to contact us for updated information in this regard.

Aspects such as the size or concentration of the product, information supplied by the producer, variety of products offered, or the absence of special transport conditions due to lyophilized presentation make of Vircell PCR controls a highly competitive line in the market. According to the nature of these kinds of products and the great variability of customers (knowledge, know-how, means, regular practice, etc) it is very difficult to determine the number of tests performed with a PCR Control. Thus, the comparison price per test should be done considering every particular protocol and customer.

For example, some manufacturers offer non-quantified PCR controls containing 100 µl at a final concentration of 50 ng/µl. That is the same relation of volume-concentration that Vircell offers in its non-quantified PCR controls. However, these manufacturers’ end-user-price almost doubles ours. Likewise, non-lyophilised PCR controls require dry ice for its transportation, which increases its cost. Moreover, the transportation costs of non-lyophilized PCR controls are much higher as they require dry-ice. Vircell PCR controls are always offered in lyophilized vials.

The PCR controls containing bacterial DNA that are in the catalogue at the moment are not provided within a capsule. Vircell PCR controls just contain the complete and purified genetic material from the bacteria ready to use in amplification reactions.

- Limited quantity available
- Difficulty to obtain positive samples in esoteric or less frequent parameters
- Non standardized results; lot-to-lot inconsistency
- Non-purified controls- human and other microorganisms DNA present in the controls
- Risks associated with infectious agents - Less stability
- Only quantified DNA/RNA controls can be used in quantitative PCR

The use of commercial PCR controls avoids these difficulties and provides advantages in terms of a friendly use, long-term storage capabilities, standardization, quality and consistency. Also, in some countries laboratories are encouraged by the accreditation groups to use third party controls.

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