1.Selection of a cellular material.

It is possible to prepare for preparations of chromosomes from all tissues and the cellular suspensions containing dividing cells: chorine, a bone marrow cells , sperm, amniotic cells. At human in most cases use preparations from cells of a bone marrow, from culture of cells of blood or from culture of fibroblasts. The most simple method is cultivation blood cells.

2. Metaphase method:

The most convenient object for these researches is a culture of lymphocyte of peripheral blood in blood of healthy people is not present dividing cells. Mitosis of these cells possible to stimulate artificial: 1-2 ml of venous blood + the mix of a nutrient medium with FGА (phytohaemagglutinin – stimulator of mitosis) causes division of lymphocytes. Suspension of leukocytes is raised in cultural environment during 48-72 hours and then prepare for preparations of chromosomes. For 2-3 hours before finishing of cultivated a division of cells is stop on the stage of metaphase, destroying of a spindle of division with help of colchicines. Chromosomes are strongly condensed and easily separated from each other. Then, the cells process a hypotonic solution (chloride of calcium or citrate sodium) for free distribution of chromosomes in a plane of a preparation. Then the cells are fix a mix methanol and an acetic acid (3:1). A drop of suspension render on glass, at drying a chromosome are strongly attached to glass and then their paint.

3.Colouring.

There are 3 groups of methods of painting: simple, differential, fluorescent

- Simple or on Gimse (« routine painting ») or 2 % acetoorcein or acetcarmin. These dyes paint chromosomes entirely, in regular intervals and intensively (centromere, satellites and secondary strangulation). This method is quite sufficient for revealing numerical anomalies of chromosomes; for studying chromosomal mutagenesis at check of factors of an environment.

With the help of this method the basic chromosomal illnesses were open, the role of chromosomal anomalies in spontaneous abortions, IVD (innate vices of the development) and carcinogenesis is shown, principles of biological dosimetry are developed.

For reception of more detailed picture of structure of chromosomes or their segments use various ways of differential coloring.

- Differential coloring is provided with simple temperature-salt influences on the fixed chromosomes. By means of this coloring we reveal the interleaving eu- and heterochromatic sites of chromosomes. New techniques for the analysis of chromosomes are developed: use of fluorescent dyes, coloring of chromosomes after special processing by paint Giemsa. These methods establish a precise differentiation of chromosomes of the person on their length on painted and not painted strips. Figure of these strips is specific, individual for each pair chromosomes. All methods reveal the same structures, but each of them is specific concerning the certain chromosomal segments. Various types of segments designate on methods with which help they come to light most.

Ways of differential coloring: Q, G, R, C.

а) Q – segments (quinacrine) are the sites of chromosomes fluorescing after coloring quinacrine – yperite. Q – segments correspond to sites rich A – Т pairs of DNA. By means of Q-method possible to identify Y- chromosome of the human as pair luminous points. For viewing it is necessary to use a luminescent microscope.

b) G – segments (Giemsa). First chromosomes are process protease (or incubate in a salt solution). This processing breaks structure of chromosomes which is restored at painting in part. That gives individual striation (it is supposed, that the painted segments – heterochromatin, the unpainted segments - euchromatin).

C) R – segments (reverse) are painted after thermal denaturizing. R – segments correspond to sites rich G – C pairs of DNA which are steadier to thermal denaturizing.

d) C – segments (constitute heterochromatin). It is a painting of perycentromere areas of chromosomes.

Achievements of molecular cytogenetics have allowed introducing in clinical cytogenetics to new technologies such as DNK-diagnostics, hybridization of nucleonic acids on a preparation, computer systems for analysis of chromosomes. DNK diagnostics is based use of technology recombination molecules DNK for revealing molecular-genetic defects of chromosomes. Fluorescent hybridization on preparation, FISH – diagnostics( Fluorescent In Situ Hybrization) includes application fluorescing DNK-tests for revealing genetic defects at a chromosomal level. Identification of a chromosomal pathology is based on use various DNA-probes allowing to mark individual chromosomes or their separate sites.

In 1960 the special commission had been authorized general system of a designation of chromosomes of the human (Denver, the USA). All chromosomes are numbered from 1 up to 22 according to their length, and also other features of their structure. Sexual chromosomes have no numbers and are designated X and Y.

Group A (1 – 3 pairs). 1 and 3 pairs are large metacentric chromosomes, 2 pair is submetacentric chromosomes. The first chromosome contains secondary strangulation in subcentromere area of one of shoulders.

Group B (4-5 pairs). Large submetacentric chromosomes.

Group C (6 – 12). Middle submetacentric chromosomes. The X – chromosome is similar to the longest chromosomes of this group with 6-th and 7-th. 6, 7, 8 and 11 pairs of chromosomes from this group are more metacentrics. 9,10,12 pairs of chromosomes are more submetacentric. The 6-n chromosome has secondary strangulation on a long shoulder.

Group D (13 – 15 pairs). Middle acrocentric chromosomes. All three pairs of chromosomes are potential satellite. The long shoulder of one of these chromosomes has secondary strangulation.

Group E (16 – 18 pairs). Short submetacentric chromosomes. The chromosome 16 – 17 contains on a long shoulder secondary strangulation.

Group F (19 – 20 pairs). Short metacentric chromosomes.

Group G (21 – 22 pairs). Very short acrocentric chromosomes. Two chromosomes on a short shoulder have satellites.

The X-chromosome belongs to group C.

The chromosome belongs to group G. It is a small acrocentric chromosome, has no satellites. It differs from chromosomes 21 and 22 to length a shoulder. In the middle of a long shoulder is present secondary strangulation.

Cardiograms or ideogram is an arrangement of chromosomes on groups in decreasing order the sizes.