Filter in situ hybridization (HISF). This method also requires DNA extraction and analysis of samples obtained by scraping or swab or brush. The cells are applied directly to a filter which is treated with alkali to cell lysis and DNA denaturation. HISF The method is very easy to implement, fast and inexpensive, however, little specific and often difficult to separate the positive signals from the bottom (1). Its use is limited to work with large numbers of specimens.

Southern blot hybridization (SB). Need for DNA extraction from the sample with phenol / chloroform and the removal of RNA and proteins of the same by enzymatic digestion. With the implementation of endonuclease-restriction enzyme Pst I is the most utilized, since the genomes of HPVs cut into fragments of different sizes offering a characteristic pattern in the agarose gel DNA-breaks, performing an electrophoretic separation of fragments agarose gel which can be visualized with ethidium bromide. The DNA is denatured and transferred to a membrane hybridization carried out. The SB is the current reference method for typing of HPVs. The specificity of this technique is superior to any other because of the extra purification of DNA by electrophoresis. He is the only method to know whether the viral genome is episomal or integrated state in cellular DNA. SB The fundamental disadvantages are its technical complexity, length of it and the need for unfixed tissues, since there are protocols for fixed material, its sensitivity decreases very markedly.

The blot invert (BI) is a variant of the SB. The cellular DNA is labeled and hybridized with different DNA-HPVs, previously digested and arranged in a filter. The IB requires an individual reaction to each specimen, lower sensitivity to SB (detected HPV 10 copies / cell), is commonly used for many tipar genotypes in one specimen.

The dot / spot blotting (DB) is the total cellular DNA extraction and distorted, then fixed to a membrane by negative pressure in a manifold apparatus. You can also perform the distortion in the membrane. Once hybridization occurs autorradigrafía if hot probes were used or disclosed enzyme, if used cold. With the DB can be analyzed at once more than 100 specimens while the SB does not support more than 15. One of the most important problems of the DB is the “background” caused by nonspecific hybridization. This problem can be minimized with the use of RNA probes because RNA-DNA hybrids have a higher Tm for DNA-DNA and nonspecific hybridization can be eliminated with the implementation of RNase (2).

The sandwich hybridization (HS) does not require DNA extraction. This technique uses a probe with two separate fragments, each with a region complementary to the target nucleic acid. A fragment is attached to the filter and sets the specific sequence of HPV to the same. The second fragment is checked and fixed the hybrid anchored in the membrane allowing their detección.La HS is a method with an estimated specificity in 1-5 x 10 5 molecules of HPV DNA (1 to 5 times more than the DB) and very sensitive (3 times the DB). The HS has several disadvantages, firstly only one HPV type per sample can be analyzed further to obtain good results requires a significant amount of sample greater than 5 X 10 5 (3).

Northern hybridization was used for RNA detcción of HPVs. The cellular RNA is extracted and separated by a denaturing gel containing formaldehyde. Probes are then hybridized with antisense RNA or DNA.

In all systems described above hybridization target nucleic acid is located on a solid support either a membrane or a biopsy or cytology in the procedures described below the nucleic acid is dissolved in a liquid.