Chemistry: electrical and wave energy – Apparatus – Electrophoretic or electro-osmotic apparatus
Reexamination Certificate
1999-08-30
2001-09-18
Warden, Jill (Department: 1743)
Chemistry: electrical and wave energy
Apparatus
Electrophoretic or electro-osmotic apparatus
C204S618000
Reexamination Certificate
active
06290831
ABSTRACT:
FIELD AND BACKGROUND OF THE INVENTION
The present invention relates to an electrophoretic assembly and system, and a method of utilizing same, which are utilizable for the real-time analysis of biopolymers contained within a plurality of samples.
Electrophoresis, an analytical separation and analysis technique, has become an indispensable tool in science, particularly in the analysis of biological samples of biopolymers such as proteins, DNA and RNA.
Electrophoresis is based on the fact that, under the effect of an electric field, molecules migrate within a substrate medium at different speeds according to their charge density, size and conformation. Under electrophoretic conditions wherein charge density and conformation are constant, electrophoretic separation of biopolymers serves for determining the size of the electrophoresed biopolymers. Such conditions characterize linear double stranded nucleic acids in substantially natural environment, denatured single stranded nucleic acids (e.g., in presence of urea, formamide, formaldehyde, glyoxal) and denatured and reduced proteins (e.g., subjected to a reducing agent such as &bgr;-mercaptoethanol and denaturing and charge density forming agent sodium dodecyl sulfate).
Under electrophoretic conditions wherein charge density and size are constant, conformational changes between electrophoresed biopolymers are detectable. This is the case, for example, for circular nucleic acid molecules which can resume one of several conformational forms, depending on the level of super coiling, and for single or double stranded nucleic acid molecules which differ in sequence by as little as a single nucleotide per several hundred bases, as is the case is single stranded conformational polymorphism (SSCP) and in heteroduplex analyses.
In special cases electrophoresis is employed under (hanging conformational or charge conditions, such as in denaturing or temperature gradient gel electrophoresis (DGGE or TGGE) of nucleic acids and isoelectric focusing of proteins. In other cases, alternating electrical fields are employed for electrophoresis in what is known as pulse field gel electrophoresis.
Electrophoresis is generally carried out on a solid substrate such as a gel, for example, a polyaciylamide or agarose gel, within which the molecules to be analyzed migrate at different speeds according to the underlying principles described above.
After a given migration period, during which the molecules are subject to the effect of an electric field, they are displaced within the substrate by a given distance which is proportional to their speed of migration which correlates to their charge density, size and/or conformation. By employing a suitable detection, the regions reached by the different types of molecules at the end of a given time period can be displayed in the form of bands.
Many variations of the technique have been developed over the past two decades, including slab and tube (e.g., capillary tube) gel electrophoresis.
Dedicated equipment for carrying out the different types of electrophoresis has been developed and is commercially available. Such equipment includes systems for native and denaturative protein gel electrophoresis, protein isoelectric focusing gel electrophoresis, nucleic acids gel electrophoresis, nucleic acid sequencing gel electrophoresis, denaturing/temperature gradient gel electrophoresis and pulse field gel electrophoresis.
Present day commercially available electrophoretic apparatuses typically utilized for analytical electrophoresis suffer from a number of limitations.
Since most of the apparatuses employed for electrophoresis utilize a single gel slab, a limited number of samples can be co-analyzed.
As such, when processing a large number of samples, multiple electrophoretic runs are often necessary which can be time consuming and tedious to execute. In addition comparisons from samples or runs and particularly from different sample batches are very difficult since conditions of the electrophoresis vary and regulation and monitoring of the conditions is not available or unreliable. Thus, when resorting to multiple electrophoretic runs, a wide variation in determinations of molecular weight, as well as the properties of the sample components can be experienced. Overcoming this limitation requires the use of a large gel slab with a large sample number capacity which in turn requires the use of a relatively large and thus cumbersome apparatus. In addition, the large gel slab of such an apparatus would require a high electrical current, which causes over heating of the slab.
In order to overcome these limitations, and yet maintain a relatively small apparatus size, multiple gel slab apparatuses have been devised. For example, U.S. Pat. No. 5,047,135 discloses a gel electrophoresis system in which the gel slab is a multi-gel slab which is subdivided by means of thin divider sheets into wafer-like micro-gels. This composite slab gel is electrophoresed as a single slab gel, following which the micro-gels may be peeled off and separately processed.
Although this system enables multiple gel slabs to be electrophoresed in a relatively compact instrument, there is no possibility of performing real time and simultaneous analysis of the results.
Currently available gel electrophoretic apparatuses including multi-gel slab apparatuses suffer from another inherent limitation. These apparatuses usually do not prevent the sample from running off the gel, nor do they provide assurance that the sample has had sufficient time for a reasonable separation. Thus, to assure optimal separation without sample loss a user must closely monitor the apparatus when used. This limitation is especially inherent to multi gel slab apparatuses since in these apparatuses a variance in the separation can often be experienced from one gel slab to the next, which variance cannot always be detected by the user due to the configuration of the apparatus.
U.S. Pat. No. 5,275,710 describes an electrophoretic system for carrying out electrophoresis while monitoring and analyzing, in real time, samples electrophoretically separated thereby. Although this system overcomes some of the limitations described hereinabove it is still limited to use with a single gel slab and therefore it does not provide a user with means with which real time analysis of a large number of electrophoresed samples can be effected.
There is thus a widely recognized need for, and it would be highly advantageous to have, a gel electrophoresis system for processing a large number of samples which system enables a real-time analysis of biopolymers contained within each sample.
SUMMARY OF THE INVENTION
According to one aspect of the present invention there is provided an electrophoretic system for real time analysis of a plurality of samples each containing at least one biopolymer, the system comprising (a) a holder for holding a plurality of removable gel cassettes, the holder supporting a plurality of electrode pairs electrically connectable to an external power source; (b) the plurality of removable gel cassettes, each having an open top end and a bottom end and being designed for containing a gel slab therein, wherein each pair of the plurality of electrode pairs serves for creating a voltage gradient in a single gel slab, such that electrophoretic separation of the at least one biopolymer of each of the plurality of samples is effectable thereby; (c) a plurality of detector assemblies, each including a plurality of detection units, each of the plurality of detector assemblies being intimately positionable at a region of one of the plurality of removable gel cassettes corresponding to a portion of the gel slab, each of the plurality of detector assemblies being for collecting information pertaining to at least one property of at least one biopolymer from the portion of the gel slab, whereas each of the plurality of detection units of a single detector assembly of the plurality of detector assemblies serves for collecting information pertaining to at least one property of at least one biopoly
Liran Yoram
Tal Michael
Integrated Genetic Devices Ltd.
Noguerola Alex
Warden Jill
LandOfFree
Electrophoretic system for real time detection of multiple... does not yet have a rating. At this time, there are no reviews or comments for this patent.
If you have personal experience with Electrophoretic system for real time detection of multiple..., we encourage you to share that experience with our LandOfFree.com community. Your opinion is very important and Electrophoretic system for real time detection of multiple... will most certainly appreciate the feedback.
Profile ID: LFUS-PAI-O-2542535