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The fixative is added to the cells and resuspended well with a plastic pasteur. Working under a chemical safety cabinet, it should be remembered that the degree of saturation of the filters is a critical factor since the duration of a filter depends not only on its particle size and on the concentration and frequency of use of the substances to be absorbed, but also from the thermo-hygrometric conditions of the environment and from the ability to absorb volatile substances present in the laboratory even during periods of non-operation of the safety cabinet, therefore their periodic replacement is essential. The tubes are housed in the cruise and the last centrifuge is always done at 2100 rpm for 5 minutes. At the end the supernatant is removed and finally the pellet is resuspended in only about 200 microliters of fixative. At this point proceed with the drop. Two specimen slides are cleaned thoroughly with alcohol and drops of suspension (cells and fixative) are dropped onto clean and cold slides. Methanol with a lower evaporation point evaporates earlier while acetic acid evaporates more slowly thus reducing the more accessible chromosome proteins, in addition to the entire cytoplasmic and nuclear matrix. The cell then deflates and flattens forces the chromosomes, which will tend to stretch and stretch, to spread on the slide within the margins of the membrane still present and thus ready for the final coloring. 5% Giemsa is used in deionized water, a specific dye for DNA. The Giemsa is filtered in order to remove any lamellas formed by the oxidation of the dye when exposed to direct light. 95 milliliters of water is poured into a Coplin jar and then 5 milliliters of filtered Giemsa is added and mixed. Giemsa staining is based on the differentiation of the cellular constituents that have a basic reaction, which fix the eosin (acid) by coloring in red-orange. The other cellular components having acid reaction are colored in blue, with the oxidation products of methylene blue (basic) blue. The slides are housed in the coplin jar and incubated in the coloring solution for 10 minutes in the dark. After 10 minutes the slides are rinsed in deionized water and left to dry under a chemical safety cabinet. The upright, for microscopy, is used once the coloring is complete, this phase is necessary to close the preparation with a cover-glass, to make the slide stable over time. On dry cells or on a very clean coverslip, a few drops of upright are dripped with a pasteur, and it is poured over the preparation, so that the upright will tend to spread, the excess of upright that comes out of the edges is pressed and removed. At this point the slide is ready for observation under an optical microscope. The microscope is an instrument that allows us to see what is normally not seen with the naked eye, using a system of biconvex lenses and relying on the phenomenon of refraction of light radiation. It consists of a mechanical part and an optical system. The first is divided into a base to allow good stability of the device, a stand that is the vertical support that holds the various parts of the microscope together and a table for objects which is the plane on which the slide is housed, finally a coarse and a micrometric screw on the sides of the stand which allow the sample to be focused. The optical system consists of an eyepiece that can be simple or double, with 10X magnification, the prism box with a prism that has the task of conveying the light beam that passes through the specimen, the objectives mounted on the revolver and the condenser located under the storage table and is used to concentrate the light beam emitted by the bulb below. After focusing, one looks for a metaphase and proceeds with its observation to make the karyotype. Chromosomes are counted: important to suspect alterations in the number, then the homologues appear by similarity, that is. length and position of the centromere. Finally, they are sorted according to the Denver Convention, proposed in 1960 at the Congress of Human Genetics held in Colorado. Cell cultures prior to centrifugation are spilled into plastic centrifuge tubes with cone bottom 24.45 milliliters of medium are collected in two stages: 24 milliliters with the pipettor using a 25 milliliters serological tip and subsequently I will withdraw 450 microliters with the single-channel pipette In the preparation of the culture medium, the complete for each soil tube requires: RPMI medium 4 point 0 75 milliliters for six tubes 24.45 total millilitres, 10% fetal bovine serum 0.5 milliliters for six tubes three milliliters total, then one percent antibiotics 10,000 units per milliliter 0.0 50 milliliter for six tubes 0.3 milliliter total, finally 1.5% of phytohaemagglutinin 0.0 75 milliliters for six tubes 0.45 milliliters total