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Cellogel is a film of cellulose
acetate in gel form, produced in wet state to maintain its gel properties
and facilitate the impregnation into the buffer solutions without
the problem of air being trapped in its pores as can occur when
using dry microporous acetate membranes. Cellogel is the ideal electrophoretic
support for clinical electrophoresis and for the immunological techniques
where it often out-performs agarose. Cellogel is an electrophoretic
medium which separates the proteins, even at high resolution, according
to the electric charge and does not have the effects of molecular
filtration typical of other gels like polyacrylamide. Cellogel is
packed in strips and sheets of various dimensions. |
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If compared with German or American
produced dry cellulose acetate (in microporous film for electrophoresis)
Cellogel presents important properties and advantages: |
| a. |
Cellogel is ready for buffering and does not entrap
air at the moment of immersion into the electrophoretic buffers. |
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| b. |
In comparison with dry acetate,
with a thickness from 120 to 160 microns, Cellogel is produced with
thicknesses between 190µ up to 500µ depending on what
it is to be used for. The greater the thickness, greater is the
volume of the specimen which can be deposited on it (for example
with Cellogel with a thickness of 300µ a semi-micro applicator
of 1.2µl/9 mm can be utilised instead of the semi-micro of
0.9µl/9 mm normally used with 200µ Cellogel; this signifies
that with specimens which are poor in proteins it is advisable to
use a thicker Cellogel, thus depositing a larger quantity of proteins
to be detected). Furthermore, higher thickness corresponds, with
the same voltage applied during electrophoresis and with the same
ionic strength of the buffer, to a higher passage of current measured
in mA x strip (e.g. a 5.7x14 cm strip of 200µ Cellogel impregnated
with a buffer with ionic strength equal to 0.05 placed on a bridge
8.5 cm long in a 200 V electric field permits the passage of 5 mA
per strip. If the same Cellogel were 250µ thick, it would
allow the passage of 6.5 mA per strip, if were 300µ it would
allow the passage of 7.5 mA and therefore (V x mA) more Watts). |
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| c. |
With Cellogel there is the possibility to apply
specimens with a volume of 0.9µl/9 mm (semimicro method) or
of 2µl/18 mm (macro method) without the sample spreading as
would occur on a very thin dry acetate strip which tolerates micro
applications of 0.25µl/4 mm well but lets the semi-micro and
macro deposits spread unacceptably. The application can be repeated
two or three times on the same spot on Cellogel, when necessary,
as in the case of electrophoresis of isoenzymes and of biological
liquids poor in proteins. |
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| d. |
Dry acetate is limited to the migrations of
20 mm of miniaturised micro electrophoresis or at most of 30 mm
with a quasi-semi-micro carried out with stamp applicators and their
relative dispocards. Cellogel, however, is suitable for standard
migrations of semi-micro 35 mm serum proteins, with 45 mm semi-micro
with prolonged migrations or high resolution electrophoresis with
60-70 mm migrations or more. |
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| e. |
HRE (high resolution electrophoresis) is only
possible on Cellogel and not on dry acetates. HRE on Cellogel is
much simpler and easier than on agarose; the expensive systems for
the circulation of cold water or Peltier control which are needed
for all the commercial agarose gels with a thickness of 500 microns
are not required with Cellogel. HRE on Cellogel has a cost per test
equal to a semi-micro test on acetate and does not have the prohibitive
costs of agarose which is only produced in kits of 10 or maximum
15 tests per film, which cannot be proposed for the routine of large
and medium size laboratories. With French agarose it is only possible
to carry out 10 tests/hour, with American agarose 15 tests/hour,
while with Cellogel it is possible to perform up to 48 test/hour;
furthermore HRE on agarose presents itself with migrations containing
a floating ß-lipoproteins fraction focused, sometimes, overlapped
on a small monoclonal band. In practice, high resolution on agarose
is a time consuming system as well as being defective. Cellogel,
like agarose, offers resolutions that depend on the length of the
migrations. Making a deposit of 0.9µl on a line 9 mm long
and 1.5 mm wide (semi-micro deposit): |
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- After 35 mm movement of albumin the serum proteins migration
shows 5-6 fractions
- After 50 mm it shows 7-9 fractions
- After 65 mm it shows 9-13 fractions
- After 110 mm it shows between 11 and 23 fractions
Chemically Cellogel is a film of water made of from 7-8% of solid
cellulose acetate and 92-93% H 2O
of which 60-70% is constitution H 2O
bound with hydrogen bridges, and 20-30% water for impregnation of
the pores. The evaporation and water transport onto the membrane
during prolonged electrophoresis is better regulated, the evaporation
of the constitution water bound by the hydrogen bridge is much slowed
down and this facilitates long migrations which are impossible on
dry acetate. The porosity of Cellogel is predisposed for the main
analysis, that is electrophoresis of the serum proteins. Large molecules
like pre-ß-lipoproteins and all the other serum proteins penetrate
and migrate. Only the chylomicrons do not penetrate or migrate and
only leave a mark at the start point, the same occurs with immunocomplexes
and cryoglobulins when present; these marks which are analytically
and diagnostically important, cannot be seen on the French agarose
which uses filtering applicators.
The predisposed porosity of Cellogel is decisive in avoiding spreading
of samples at the moment of depositing and spreading of the fractions
with low mobility during migrations which can be lengthy. All in
all the right porosity corrects the insufficiencies of other commercial
cellulose acetates membranes. To this must be added the better compatibility
between Cellogel and serum proteins, including lipoproteins, that
are incompatible with agarose. The latter is, in fact, a film of
water (99% H 2O) totally hydrophilic,
where the amphiphilic serum proteins with more lipophilic characteristics
remain floating on the surface even when the sample is deposited
with applicators which cut the gel.
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The superiority
of Cellogel over agarose was recognised in numerous publications
by important authors between 1963 and 1971. Thanks to its amphiphilic
properties (hydrophilic and lipophilic) Cellogel has optimal compatability
with specimens as difficult and complex as serum proteins, which
are also amphiphilic. Cellogel is, therefore, the ideal support
for electrophoresis of serum proteins, hemoglobins, lipoproteins,
isoenzymes, for all the immuno-electrophoretic techniques and for
the search for antigens, antibodies and tumour markers (especially
those immunofixable with polyclonal antibodies). |
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