Chemistry: electrical and wave energy – Processes and products – Electrophoresis or electro-osmosis processes and electrolyte...
Patent
1995-03-13
1998-11-24
Gorgos, Kathryn L.
Chemistry: electrical and wave energy
Processes and products
Electrophoresis or electro-osmosis processes and electrolyte...
204464, 204613, 204614, G01N 2726
Patent
active
058401695
DESCRIPTION:
BRIEF SUMMARY
This application is a 371 of PCT/DK92/00058 filed Feb. 25, 1992.
FIELD OF THE INVENTION
The present invention concerns a process and apparatus for electroelution of a gel containing charged macromolecules, such as proteins or DNA/RNA.
BACKGROUND
Electrophoresis of proteins or DNA/RNA in gels is widely used to separate compounds which differ in size and/or charge. Such compounds exhibit different migration distances and can be investigated by staining the gel or by transfer to a membrane suited for the study of antibody-antigen or other affinity interactions. Recovery of individual protein bands, however, is a laborious and time consuming procedure which generally has been based on cutting the gel into segments containing a single band, followed by homogenizing and elution, either by electroelution or by simple diffusion. Such a process is often ineffective and yields very high dilutions.
U.S. Pat. No. 3,956,099 discloses a complex system suited for continuous operations. The system separates components into separate compartments during the electrophoretic run. The procedure is not based on the elution of macromolecules already separated in a conventional gel, but includes instead a separating unit suited for large scale purification of proteins.
The processes and components described in U.S. Pat. Nos. 4,049,534, 4,725,348 and 4,747,918 all suited for electroelution of small gel fragments containing single components. The procedures include a preceeding localization and excision of the desired molecule, a technique which is time consuming and involves problems with respect to accuracy. These methods are therefore not suited for the simultaneous investigation of all components separated in a gel.
EP Published Application No. 0 380 313 discloses a field inversion electroblotting and elution device specially suited for transfer of high molecular mass compounds. The procedure includes a preceeding identification of the band of interest and, therefore, has the same limitations as mentioned in relation to the methods described in the above-mentioned U.S. Pat. Nos. 4,049,534, 4,725,348 and 4,747,918.
U.S. Pat. No. 4,181,594 discloses a matrix recovery apparatus wherein compounds contained in a slab gel are electroeluted directly upwards into multiple wells. Each compound is eluted into several wells separated by walls of a considerable thickness.
The Blotelutor.RTM. apparatus (manufactured by Biometra, Gottingen, Germany) provides a unit which is exclusively suited for the elution of proteins from two-dimensional gels. The proteins are eluted into wells separated by walls of a considerable thickness. Further, the method underlying this apparatus is not based on a physiological buffer and does not include a pre-equilibration step to ensure a fixed positioning of the gel during elution.
None of these prior systems meet the need for conventional, simultaneous elution of all compounds separated in a one-dimensional gel. A system with these features would greatly facilitate the biological and chemical screening procedure of complex protein or polynucleotide mixtures.
SUMMARY
It is an object of the present invention to provide a process for simultaneous elution of separated charged macromolecules such as DNA/RNA contained in a one dimensional gel, and to provide an apparatus and method of use in carrying out the process. According to the process of the invention, a simultaneous elution of a mixture the separated charged macromolecules, such as proteins, in an entire gel, such as a polyacrylamide or agarose gel, is performed. The charged molecules are, thus, divided into narrow fractions each containing single or closely located compounds, and the greater part of ione detergents, such as sodium dodecyl sulphate (SDS), as is removed.
The novelty according to the process of the invention is the collective elution of an entire gel. The present invention additionally provides an electroelution unit suited for the simultaneous elution of all compounds separated in a whole gel by the process of the invention. The unit
REFERENCES:
patent: 3956099 (1976-05-01), Israel et al.
patent: 4049534 (1977-09-01), Posner
patent: 4181594 (1980-01-01), Rizk et al.
patent: 4608147 (1986-08-01), Clad
patent: 4725348 (1988-02-01), Diekmann
patent: 4747918 (1988-05-01), Wassenberg, II
patent: 4775636 (1988-10-01), Moeremans et al.
patent: 4948481 (1990-08-01), Miillner
A. T. Andrews, Electrophoresis: Theory, techniques, and Biochemical and Clinical Applications 2nd Ed., Clarendon Press-Oxford, 1986.
Anderson Denton L.
Farah David A.
Gorgos Kathryn L.
Noguerda Alex
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