Ultramicroscopy.
It is a method of detection of particles, which sizes lay outside the permitting limit of a microscope. The microscope working on this method is called ultramicroscope. In them lateral (slanting) illumination owing to what submicroscopic particles are visible as light points on a dark background are carried out.
Electronic microscopy.
For
research of structure of membranes the electronic microscope which
permitting limit is defined by length of a wave of de Broil for
moving with high speed of electron is used:
.
Where h -
Planck's constant; m - weight of electron;
- speed of electron. The permitting limit of
an
electronic microscope can be
.
Tungsten, osmium and some other chemical elements well absorb and
disperses electrons. The membrane becomes covered by these substances
for reception precise electron gram of the cells.
For research of membranes methods of freezing-breaking off and freezing-etchings are used. Preparations are quickly frozen, without damaging influences, as at reception of thin cuts. Preparation of a preparation includes a number of operations. After freezing the sample. representing suspension of cells, breaks away by means of a knife at low temperature (-1000 С) in deep vacuum. At breaking off the cut which is passing through the sample is formed. If the plane of a cut passes through a membrane, it breaks up on median area and is split on two half. The internal area of a membrane is naked on the formed planes of cut.
If necessary the sample is exposed to etching, sublimation of an ice is spent in vacuum. It allows to see superficial structures of cellular membranes better. After that the sample from the naked surface is received. This sample is studied by a method of electronic microscopy. First the sample becomes covered by platinum under a corner nearby 450 to reveal topological characteristics of a preparation. Then the sample becomes stronger, putting on it a layer of carbon. After that the preparation is thaw, the sample emerges.
Method of diffraction of X-rays radiation.
X-ray
radiation is used for the analysis of membrane structures. The main
condition of display of diffraction is comparability of the sizes of
object on which radiation goes, and lengths of a wave of this
radiation. For the analysis of small objects radiation with length of
a wave a little nanometers is necessary. A range of x-ray lengths of
its waves from 10-5
up to 80 nanometers.
Let's
consider crystal structure. Parallel beams fall on two next atoms
(fig. 4). They are reflected from the next atoms of a crystal
lattice, a
nd
then there is interference, gathering in one point in some screen.
Difference of a course of beams
,
d - internuclear distance,
- corner of sliding.
Two waves
will strengthen each other if difference of a course it is equal to
an integer of lengths of waves
Observing a picture of diffraction on the screen, it is possible to establish, under what corner the maximum of the first order (n=1) is observed, for Fig. 4. Diffraction of X-rays.
example.
Knowing length of a wave, it is possible to find the major parameter
of structure of investigated substance - internuclear distance.
By means of the X-rays t analysis have been confirmed double layer arrangement of phospholipids and presence at membranes of protein, the important structural parameters of a membrane are calculated.