Conductimetric bioassay techniques

Claim 2

A conductimetric sensor for the quantitative bioassay of an analyte in a liquid sample comprising a sensing member whose conductivity, when in contact with the sample, is proportional to the concentration of the analyte, in the sample, and a pair of spaced-apart electrodes in electrical contact with the sensing member and connectable to means for measuring the conductivity of the sensing member, wherein said sensing member comprises an essentially non conducting porous membrane with which the electrodes are in face-to-face electrical contact at two separate points spaced along the length the membrane, said membrane being formed from a material having a conductivity of less than 10 -9 Siemens/cm and having immobilised thereon a substance having a specific affinity for, or reactivity with the analyte, and reactive therewith in the presence of the sample to induce a change of conductivity along the length of the membrane between the two electrodes.

Claim 3

A sensor according to Claim 1, wherein the two electrodes are in face-to-face contact with the same face of the membrane.

Claim 4

A sensor according to Claim 1 or 2, wherein the membrane is cellulose acetate or cellulose nitrate.

Claim 5

A sensor according to Claim 2 or 3, wherein the two electrodes are embedded in an elongated insulating support adjacent one end thereof, each electrode having an exposed surface substantially flush with the surface of the support and in electrical conatact with said membrane positioned or positionable on the surface of said support in overlapping relationship with the exposed electrode surfaces.

Claim 6

A sensor according to Claim 4, wherein the membrane is carried by a slide member slidably and removably mounted on the elongated insulating support.

Claim 7

A sensor according to Claim 5, wherein the membrane is exposed for contact with the liquid medium containing the analyte to be determined through an opening in the slide member.

Claim 8

A sensor according to any one of Claims 1 to 6, wherein the substance immobilised onto the membrane is an enzyme.

Claim 9

A sensor according to Claim 7, wherein the enzyme is a penicillinase, urease, glucose oxidase, creatinase, creatininase, paracetamolase, sarcosine hydrogenase, pronase or restriction endonuclease.

Claim 10

A sensor according to any one of Claims 1 to 8, including a second pair of spaced apart reference electrodes and a second membrane positioned or positionable therebetween and in face-to-face contact therewith, said second membrane being substantially identical with the first membrane, and likewise introducable onto or contactable with said sample, but not having the said substance immobilised thereon, the said second membrane thus providing a reference value against which variations in-the conductivity of the first membrane carrying the immobilised substance, when in contact with said sample, can be measured.

Claim 11

A conductimetric bioassay apparatus comprising a sensor according to any one of Claims 1-9, and means for measuring the conductivity of the membrane(s) when in contact with the sample.

Claim 12

Apparatus according to Claim 10, wherein the sensor is as claimed in Claim 9, and wherein the measuring means comprise means for alternately measuring the conductivity of the membrane carrying the immobilised substance and the conductivity of the reference membrane, means for comparing the two values and thereby to produce a differential value, means for storing and adding successive differential values obtained over a period of time, and means for displaying a cumulative total at the end of said period, such cumulative total thereby indicating the concentration of the analyte in the medium.

Claim 13

A conductimetric bioassay method for the determination of an analyte in a liquid sample, which comprises contacting the sample with a sensor comprising an essentially non-conducting porous membrane formed from a material having a conductivity of less than 10 -9 Siemens/cm and having immobilised thereon a substance having a specific affinity for a reactivity with the analyte, the interaction between the analyte and said substance in the presence of said sample being effective to cause a change in conductivity in the membrane, and measuring the change, if any, of conductivity of said membrane when in contact with said sample, said measurement being, made along the length of the membrane.

Claim 14

A conductimetric bioassay method for the determination of an analyte in a liquid sample, which comprises contacting the sample with a sensor comprising an essentially non-conducting porous membrane formed from a material having a conductivity of less than 10 -9 Siemens/cm and having immobilised thereon a substance having a specific affinity for a reactivity with the analyte, the interaction between the analyte and said substance in the presence of said sample being effective to cause a change in conductivity in the membrane, and measuring the change, if any, of conductivity of said membrane when in contact with said sample, said measurement being, made along the length of the membrane.

Claim 15

A method according to Claim 12, wherein the membrane is of cellulose acetate or cellulose nitrate.

Claim 16

A method according to Claim 12 or 13, wherein the analyte is an enzyme substrate and the immobilised substance is an enzyme having a specific activity for that substrate.

Claim 17

A method according to Claim 14, wherein the analyte is a penicillin, glucose, urea, paracetamol, creatin, creatinin, sarcosine or nucleic acid, and the enzyme is, correspondingly, a penicillinase, glucose oxidase, urease, paracetamolase, creatinase, creatininase, sarcosine hydrogenase or restriction endonuclease.

Claim 21

A method according to Claim 18, wherein the conductivity of the two membranes are measured alternately, and a final value displayed corresponding to the cumulative total of the successive differences in conductivity between the two membranes measured over a predetermined period of time.

Claim 22

. A method according to Claim 12, as applied to a competitive assay for the determination of a ligand or a specific binding partner of a ligand present in a sample in unknown quantity, which method comprises: A) contacting the sample with a porous membrane composed of a material having a conductivity of less than 10 -9 Siemens/cm and having immobilized thereon a quantity of a specific binding partner of the ligand to be assayed, or of the ligand corresponding to the specific binding partner to be assayed, as the case may be, in the presence of a known quantity of the unknown ligand or specific binding partner, as the case may be, labelled with an enzyme, the quantity of one or other the immobilized component or the enzyme labelled component being restricted whereby competition takes place between the unknown and the enzyme labelled component for conjugation with the immobilized component; B) contacting the membrane obtained in step A and comprising an unknown quantity of the enzyme-labelled component conjugated to the immobilized component of the membrane with a substrate for the enzyme, the enzyme and substrate being such that operation of the enzyme on the substrate effects a change in conductivity of the conductance member constituted by the membrane and the immobilised enzyme-conjugate; C) measuring the change - of conductivity of said conductance member, and D) determining therefrom the concentration of the unknown in the original sample.

Claim 23

A conductimetric bioassay kit for the conductimetric assay of a ligand or a specific binding partner of a ligand in a sample, said kit comprising as a first item a porous membrane composed of a material having a conductivity of less than 10 -9 Siemens/cm and having immobilized thereon a given quantity of a ligand or a specific binding partner of a ligand corresponding to the unknown, and as a second item, a given quantity of the unknown ligand or specific binding partner as the case may be, labelled with an enzyme, being such that when conjugated to the membrane by binding with the immobilized component, and contacted with an enzyme substrate, the conductivity of the membrane member formed by the membrane and the immobilised enzyme conjugate changes as a result of the enzyme/substrate interaction and proportional to the concentration of the unknown.

Claim 24

A conductimetric bioassay kit for the conductimetric assay of a ligand or a specific binding partner of a ligand in a sample, said kit comprising as a first item a porous membrane composed of a material having a conductivity of less than 10 -9 Siemens/cm and having immobilized thereon a given quantity of a ligand or a specific binding partner of a ligand corresponding to the unknown, and as a second item, a given quantity of the unknown ligand or specific binding partner as the case may be, labelled with an enzyme, being such that when conjugated to the membrane by binding with the immobilized component, and contacted with an enzyme substrate, the conductivity of the membrane member formed by the membrane and the immobilised enzyme conjugate changes as a result of the enzyme/substrate interaction and proportional to the concentration of the unknown.

Claim 25

A kit according to Claim 22, including, as a third item, a quantity of substrate for enzyme.

Claim 26

A kit according to Claim 21 or 22, including, as a fourth item, means for measuring the electrical conductivity of the first item.