Chemistry: electrical and wave energy – Processes and products – Electrostatic field or electrical discharge
Patent
1993-01-22
1994-09-20
Niebling, John
Chemistry: electrical and wave energy
Processes and products
Electrostatic field or electrical discharge
2041821, 2041828, 2041829, 2041832, 2041833, G01N 2726, G01N 27447
Patent
active
053486331
ABSTRACT:
A method for quantitative detection of trace amounts of an analyte in a sample is disclosed. The method in the preferred embodiment includes providing an Fab' fragment of an immunoglobulin labelled at a reactive sulfhydryl group with a fluorescent dye; combining the labelled Fab' fragment with a sample that may contain the analyte; concentrating the elements of the resulting mixture in an electric field; separating the labelled analyte/agent complex formed from any unreacted labelled agent using capillary electrophoretic methods; and detecting the fluorescent signal of the labelled analyte/agent complex. The invention also is directed to a method of producing a labelled Fab' fragment that includes cleaving an immunoglobulin molecule to obtain one F(ab').sub.2 fragment; reducing the disulfide-bonds of the F(ab').sub.2 fragment to obtain two Fab' fragments each having at least one free, reactive sulfhydryl group; and mixing an Fab' fragment having at least one free sulfhydryl group with a fluorescent dye reactive with the free sulfhydryl to form a labelled Fab' fragment. Preferably, prior to the final mixing step, intrastrand disulfide bonds are formed by oxidation within each Fab' fragment, thereby producing individual Fab' fragments each having a single reactive sulfhydryl group. The method of quantitative detection also more broadly includes using any biospecific agent to form a complex with the target analyte.
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Karger Barry L.
Shimura Kiyohito
Niebling John
Northeastern University
Starsiak Jr. John S.
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