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CITOMEGALOVIRUS VIRAL LOAD

Categoría: Infectious diseases.
Días hábiles de entrega: 7
 

EDTA blood, urine, LCR.

Store samples at temperatures between 4-8 ° C, with refrigerant gels or ice pack.

DNA extraction from the sample using commercial kits. Real-time PCR using primers and the TaqMan probe for the quantification of CMV DNA that were designed from the phosphorylated matrix protein gene pp65 and synthesized by BioSource Int. According to Griscelli F, et al. 2001. DNA amplification is detected as an increase in fluorescence. A Chromo4 Four-Color Real-Time System thermocycler (MJ Research, MA, USA) is used for the realization of the technique. Positive control: ACCURUN 350 (CMV DNA Positive Control Series 300, BBI Diagnostics, MA, EU) with a known copy number and CMV AD169 strain (Ref ATCC).

10 working days.

Result: CMV DNA NOT DETECTED LOG: --

Result: CMV DNA DETECTED, ______ Copies /ml LOG: ___

Result: CMV DNA DETECTED,

Detection test limit: 100 Copies / ml

The quantification of CMV by real-time PCR shows the usefulness of this test in the monitoring of changes in CMV DNA levels. This technique may be the best alternative for the diagnosis of infection, for predicting diseases and for monitoring the effectiveness of antiviral treatment.

It is important to note that monitoring a patient's viral load should be done with the same methodology and by the same laboratory. This guarantees uniformity and reliability in the results taking into account that the use of different techniques complicates the interpretation of the results. On the other hand, viral load levels may not always be comparable between the different techniques since there may be variations due to other factors such as differences in samples, nucleic acid extraction methods, primers, patterns and detection methods.

It is recommended that absolute values of viral load obtained by genomic amplification procedures be interpreted as any value within a range of 0.5 log (± 300 times) of the value obtained.

This test has been validated by Genomik Laboratory with positive and negative commercial controls; however, a negative result does not exclude the presence of CMV DNA below the sensitivity limit of the test.

Cytomegalovirus (CMV) infection is very frequent and generally asymptomatic. However, the incidence and clinical conditions it produces in newborns and immunocompromised patients make this virus an important human pathogen. It can cause severe illness and death in immunocompromised individuals, including bone marrow transplant recipients (BMT), solid organ transplant recipients, patients infected with human immunodeficiency virus (HIV), patients taking immunosuppressive drugs, and newborns infected during the pregnancy. In children, it is the most common cause of congenital and perinatal infection, affecting between 0.4% and 2.3% of newborns and in immunosuppressed patients is the major cause of morbidity and mortality. These patients at high risk of fatal CMV disease need early, sensitive and rapid diagnosis. In addition, in many cases the effectiveness of antiviral treatment depends on the detection of CMV viral load in the early stages of infection. Quantitative PCR tests for CMV DNA have been shown to be useful for rapid diagnosis of infection and for effective monitoring of clinical evolution in response to treatment. In recent years, real-time PCR has been widely used to determine viral load during CMV infection.

Griscelli F, Barrois M, Chauvin S, Lastere S, Bekket D, Bourhis JH. 2001. Quantification of human cytomegalovirus DNA in bone marrow transplant recipients by real-time PCR. J Clin Microbiol, 39: 4362-69.

 



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