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Electrophoretic method for the isolation and separation of microorganisms and cell populations
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Inventors: Ebersole, Richard C.; DeCarolis, John G.; McCormick, Randy M.;
Assignee: E. I. Du Pont de Nemours and Company (Wilmington, DE)
Primary Examiner: Gitomer; Ralph J.
Assistant Examiner:
Attorney, Agent or Firm:
The present invention sorts microorganism populations from a mixture which contains more than one microorganism population. Microorganisms of different types vary in size, shape, and surface charge characteristics. These characteristics we believe contribute to the rate of migration for microorganisms under the influence of an electric field. Microorganisms of the same population (genus and species) will migrate similarly. By applying an electric field in a direction opposite the direction of fluid flow, separation is enhanced. |
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DETAILED DESCRIPTION It is an object of the invention to provide a novel electrophoretic method for the separation of viable mixtures of viable bacteria for analysis of microbial communities.
The instant method utilizes a small bore capillary tube filled with a suitable electrolyte and the counteracting forces of both an electrophoretic field and an electroosmotic flow to effect a highly specific separation of bacterial and other biological cells.
Furthermore, it is also an objective to provide a method to isolate and collect the separated bacterial fractions, using a computerized automated robotics collection device.
The invention further provides an apparatus and a means to dispense the separated bacterial fractions by streaking them onto the surface of a nutrient agar gel plate, in this way providing a biogram of cell composition and cell numbers.
The invention also provides means of collection in which the bacterial fractions can be dispensed into tubes, multi-well devices or deposited onto planar supports and membranes, which would allow further biochemical analysis, such as identification, microscopy, immunoassays, DNA assays, etc.
Sorting of the fractions can also be accomplished by using multiple detectors, which can discriminate a particular cell type having a specific detector or electrophoretic response, then transported automatically to a specified location or collection vessel.
A sample of a bacterial mixture is introduced into the anodic end of the capillary tube, either by siphoning or electrokinetic injection, that has been filled with an electrically conductive solution.
As an electric field is applied to the capillary, both electroosmotic and electrophoretic movements are created, resulting in the separation of the viable bacteria based on differences in their charge and mass.
Unexpectedly and as a result of the preponderance of the electroosmotic field, the bacteria migrate backward toward the anode while the bulk electrolyte flow is toward the cathode, and the net movement of buffer and separated bacteria is toward the cathode
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