Chemistry: molecular biology and microbiology – Animal cell – per se ; composition thereof; process of... – Rodent cell – per se
Patent
1998-02-26
1999-08-10
Witz, Jean C.
Chemistry: molecular biology and microbiology
Animal cell, per se ; composition thereof; process of...
Rodent cell, per se
435374, 435325, C12N 506, C12N 508, C12N 500
Patent
active
059358556
DESCRIPTION:
BRIEF SUMMARY
TECHNICAL FIELD
This invention concerns a method for the preparation and conservation of cellular cultures or parts of cells in the ready state for biological test.
The method according to the invention is employed in the biological, medical and research fields for the preparation and conservation of cellular cultures or parts of cells which are used as such thereafter for the performance of biological test, whereby the cells have to be further sensitized beforehand in relation to a particular hormone or a particular biological or pathological effector.
The cellular cultures or parts of cells prepared with the method according to the invention can be used also in centers not equipped for maintaining cellular cultures for the performance of biological tests, namely the so-called bio-assays.
In particular, the method according to the invention is used to prepare FRTL5 cell cultures, that is to say cells of rat thyroid, which can be employed for the subsequent monitoring and quantifying of the bioactivity of the TSH (hormone of growth) or of the thyroid-stimulating auto-antibodies (TSab).
BACKGROUND OF THE INVENTION
Biological tests are used at the present times in the medical, clinical and research fields to monitor and quantify the biological activity of activating or inhibiting factors which cannot be otherwise identified and quantified reliably with other laboratory techniques.
To carry out such biological tests, it is necessary to use animals or cellular cultures and, in the latter case, to subject the cells at times to a step of preliminary preparation called the "starvation" or "privation" step, in which the cells are bred in complete absence of a hormone, which is then used as a stimulator to increase the sensitivity of the cells to that hormone.
The cells which have undergone the "starvation" or "privation" step are called "cells in the ready state".
The sensitization is also effective in the same manner as regards pathological stimulators such as the thyroid-stimulating auto-antibodies (TSab).
This "privation" step has a duration which can be varied according to the type of tests and which lasts at least for some days, generally between about 5 and 10 days.
A first method arranges to keep the colonies of cells continually in incubation until the moment of performance of the test.
This method entails a plurality of drawbacks such as the high cost arising from the consumption of the culture medium required for maintaining cells always ready for possible tests, the danger of contamination of the culture medium with a resulting lack of validity of the tests carried out, the need to ensure the maintaining of sterility during the whole period of starvation, the need to change the medium, the maintaining of standard conditions during the whole period and the difficulty of transferring the cultures to a distance.
Moreover, this method of extempore production of the cultures when required does not ensure a production of homogeneous and reproducible cultures with the consequence that the results achieved with cultures produced at different periods and/or at different places cannot always be completely compared to each other.
So as to avoid some of these problems, a method is sometimes used whereby the cultures are frozen before the starvation step.
This second method, on the one hand, reduces the costs of the additives required during the cultivation of the cultures but, on the other hand, does not solve the problems linked to the long times of the starvation period and to the standardization of the final product.
In particular, this method consists in freezing at a temperature of at least -80.degree. C., but typically at temperatures of about -196.degree. C. in liquid nitrogen, the cells which have to be used.
These frozen cells, so as to be used, have to be subjected to thawing and to a successive period of recovery during which the cells are deposited on, and possible adhere to, the plastic bottom of the container and then undergo the starvation step.
This period of recovery may last even for some days
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Ambesi Impiobato Francesco Saverio
Degrassi Alberto
Formisano Silvestro
Lavaroni Stefano
Consorzio Di Ricerche Biomediche
Hanley Susan
Witz Jean C.
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