DNA-PCR

This laboratory method will be available from 01.02.2016!

The PCR (Polymerase Chain Reaction) is a molecular biological method for the direct detection of a pathogen’s nucleic acids (DNA/RNA) in a sample. The detectability of pathogen-DNA/RNA in blood or tissue indicates the (temporary) presence of the pathogen. A positive PCR result therefore provides direct proof of the presence of an infection with the detected pathogen.

However, it should be noted that the presence of a pathogen in a sample and thereby the detectability of pathogen-specific nucleic acids depends on the pathomechanism and also varies with the stage of disease.

 

Prior to the actual PCR-Test the entire DNA/RNA is isolated from a patient’s blood or tissue sample. These nucleic acids are then used as starting material for the requested pathogen-specific PCR-Test.

During the PCR reaction specific oligonucleotides (primer) are used for each pathogen which only bind to a sequences of a gene of the pathogen enabling amplification of this gene and subsequent detection of the pathogen.

Thus a very high specificity is achieved. Advanced technique, i.e. the nested PCR, real-time PCR or qPCR, achieve the highest sensitivity for the detection of pathogens.

 

For the detection of Lyme disease pathogens we developed our own in-house PCR testing system, which is based on multiple consecutive PCR reactions and which detects different pathogen genes with high sensitivity and specificity. The BCA-lab is therefore one of the few laboratories that is able to detect Borrelia miyamotoi. The spectrum of molecular biological pathogen detection is completed by a PCR-based Bartonella-Assay.

 

Material for blood analysis: 4 x EDTA-tubes, 1 x serum tube and 1 x CPDA-tube

Material for tissue/biopsy analysis: 1-2 pieces of tissue (> 1mm³) in sterile tubes filled with 0,9% sodium chloride solution at 4°C; for a longer transport (> 24 hr) immediately freeze tissue samples in sterile tubes without liquid at -20°C or -80°C

 

Turnaround time: 3 days (Note: Taking into account the required pre- and post-analytical work, the report is created in about 2 weeks)

 

Further reading:

Eshoo, M. W., Schutzer, S. E., Crowder, C. D., Carolan, H. E. & Ecker, D. J. Achieving molecular diagnostics for Lyme disease. Expert Rev. Mol. Diagn. 13, 875–83 (2013).

Eshoo, M. W. et al. Direct molecular detection and genotyping of Borrelia burgdorferi from whole blood of patients with early Lyme disease. PLoS One 7, 3–8 (2012).

Chan, K., Marras, S. a E. & Parveen, N. Sensitive multiplex PCR assay to differentiate Lyme spirochetes and emerging pathogens Anaplasma phagocytophilum and Babesia microti. BMC Microbiol. 13, 295 (2013).

Brettschneider, S., Bruckbauer, H., Klugbauer, N. & Hofmann, H. Diagnostic value of PCR for detection of Borrelia burgdorferi in skin biopsy and urine samples from patients with skin borreliosis. J. Clin. Microbiol. 36, 2658–65 (1998).

Cerar, T., Ružić-Sabljić, E., Glinšek, U., Zore, a. & Strle, F. Comparison of PCR methods and culture for the detection of Borrelia spp. in patients with erythema migrans. Clin. Microbiol. Infect. 14, 653–658 (2008).