CENBIMO S.L

Cytomegalovirus belongs to the β-herpes virus family and globally affects the world population.  Primary infection is often a flu-like process although there are known cases of greater severity that are clinically similar to mononucleosis.

Like other herpes viruses, after primary infection CMV persists in the host permanently as a latent infection.

Immunosuppression in patients can provoke reactivation of the virus giving rise to serious lesions in a multitude of organs such as the lungs, kidneys, central nervous system, gastrointestinal apparatus, endothelium and blood cells.

Patients with ulcerative colitis can be infected by CMV and treatment guidelines modified by this situation.

After infection, CMV first begins to transcribe the so-called ‘early’ genes.  Of these genes, one known as β2.7 (2.7 kbases) is the most abundantly transcribed and accumulates during the entire infection until it makes up 20% of all mRNA transcribed by the virus.  Because of this, the gene β2.7 is an ideal candidate for CMV identification.

CENBIMO has fabricated HISTOSONDA CMV which consists of a single strand of DNA with a sequence length of 288 nucleotides.  This sequence is complementary to CMV gene β2.7 mRNA.  The probe DNA has been labeled with digoxigenin.

The technique of in situ hybridization presents numerous advantages over immunohistochemistry and unspecific results obtained when using antibodies can be avoided.

INTENDED USE

For Research Use Only.

The Kit Histosonda Cytomegalovirus is intended for the detection of cells infected by CMV in any location found including: lymph node, central nervous system, retina, lung and intestine. This detection is carried out by in situ hybridization in paraffin-embedded, formalin-fixed histological sections.

WARNINGS AND PRECAUTIONS

The Kit Histosonda Cytomegalovirus has been designed for professional research use and must be manipulated by qualified and accordingly trained personnel.

In order to obtain the best results, the instructions contained in the manual must be followed. Any change to the indicated temperatures, times or any other step of the process can lead to poor results.

KIT COMPONENTS

The Kit includes 20 single test tubes of Histosonda Cytomegalovirus; 20 single test tubes of Proteinase K and 2 tubes of Anti-Digoxin antibody sufficient for 20 tests in total.  All products are lyophilized.

Histosonda Cytomegalovirus consists of a fragment of single stranded DNA of 288 nucleotides, complementary to CMV gene β2.7 mRNA. The probe DNA contains nucleotides that have been labeled with Digoxigenin.

STORAGE CONDITIONS

Supplied reagents are stored at room temperature until their expiration date. After being reconstituted the probes will remain stable for two weeks at 4ºC in a DNAase-free environment.  The Anti-Digoxin can be stored at 4ºC for 1 month and the Proteinase K must be used immediately and cannot be stored.  Do not use these products after their expiration dates.

SAMPLES

Any paraffin block section in which Cytomegalovirus mRNA presence is to be studied. Sections of 4-6 micrometers in width are sufficient to conduct the study. Preferably, the cut should be recent (no more than thirty days old). Assay results are not affected by block age. Studies have been carried out in the manufacturer's laboratories using 20 year old paraffin blocks with optimal results.

INTERPRETATION OF RESULTS

Samples in which Cytomegalovirus expression is observed will show a brownish color in the cell nucleus, cytoplasm or both, which will contrast over the blue-violet background given by hematoxylin staining. The pathologist will evaluate the results according to their experience, drawing conclusions from the staining of the sample in parallel with the staining observed in positive and negative controls.

ASSAY LIMITATIONS

The Kit Histosonda Cytomegalovirus has been optimized to detect Cytomegalovirus mRNA expression in formalin-fixed, paraffin-embedded tissues. Its use is not recommended for other types of samples or preparation techniques. The correct operation of this kit has been validated using the protocols indicated in the instructions manual. The use of other procedures or modification of the recommended protocols may lead to erroneous results.

The results from this assay must be evaluated by the pathologist in combination with the rest of the available patient clinical data.

In order to obtain optimal and reproducible results it is important to rigorously maintain the time and temperature conditions indicated in the procedure.